Combination lactic acid and ketobody esters for medical

Abstract

The present invention relates to a lactic acid / ketoester for protecting important organ functions and combating inflammation and cancer growth. In particular, the present invention relates to a lactic acid / ketoester having beneficial properties of lactic acid and beta-hydroxybutyric acid (BHB) for important organ functions, inflammation and cancer growth, but without the detrimental sodium load after administration of lactic acid and beta-hydroxybutyric acid.

Description

Technical Field

[0001] The present invention relates to lactate / ketone body esters for protecting vital organ function and combating inflammation, cancer growth, sarcopenia, atherosclerosis, heart failure, obesity, diabetes and metabolic syndrome. In particular, the present invention relates to lactate / ketone body esters having the beneficial properties of lactate or lactate and ketone bodies such as beta-hydroxybutyrate (BHB) on vital organ function, inflammation, cancer growth, obesity, diabetes and metabolic syndrome, but without the harmful ion loads such as calcium, potassium, magnesium and / or sodium loads that follow administration of both lactate and beta-hydroxybutyrate. Background Art

[0002] Lactate (C3H6O3) and ketone bodies, especially beta-hydroxybutyrate (BHB-C4H8O3), are both low molecular weight carbon fuel metabolites with many similarities: Both are highly O2-efficient substances that the body produces and uses as an energy source (1,2,8). They have protective properties during stress and act through similar but distinct tissue receptors (1,2). Lactate and ketone bodies contribute to the health-promoting effects of exercise, intermittent fasting, ketogenic diets, and SGLT-2 inhibition in diabetes. In addition, preclinical studies strongly suggest that both compounds (alone or in combination) are effective against inflammation, cancer growth, protein loss, and neurodegeneration (1,2). We recently showed that BHB has significant beneficial clinical effects on the heart, brain, skeletal muscle, and bone marrow (increased Epo) in terms of increased blood flow and improved organ function (3-5). The possible beneficial effects of lactate are currently being evaluated in human studies, and lactated Ringer's (sodium lactate) is routinely used in the emergency department. Several preclinical studies have shown that lactate has a variety of beneficial effects (1,2,6-9). This also includes the formation of lactic acid derivatives, such as N-lactoylphenylalanine (Lac-Phe), which suppress appetite and thus may be beneficial in patients with high BMI and / or diabetes.

[0003] WO04 / 105742 teaches compounds which reduce the level of free fatty acids circulating in the plasma of a subject and which may be used, for example, to treat muscle damage or fatigue.

[0004] US2011 / 0237666A1 discloses 3-hydroxybutyl 3-hydroxybutyrate, which is enriched in the enantiomer of (3R)-hydroxybutyl (3R)-hydroxybutyrate, as an oral precursor of (3R)-hydroxybutyric acid, for treating conditions caused by, aggravated by, or associated with elevated plasma levels of free fatty acids in human or animal subjects; cognitive dysfunction, neurodegenerative diseases, such as Alzheimer's disease, Parkinson's disease, Huntington's disease, epilepsy; hypoxic conditions, such as angina pectoris, extreme physical exertion, intermittent claudication, hypoxia, stroke, and myocardial infarction; insulin resistance conditions, such as infection, stress, obesity, diabetes, metabolic syndrome, and heart failure; inflammatory conditions including infection and autoimmune diseases, as well as muscle damage, fatigue, and muscle fatigue. It also discloses that the compound reduces the plasma level of fatty acids and can be used, for example, to treat conditions caused by, aggravated by, or associated with elevated plasma levels of free fatty acids in human or animal subjects.

[0005] WO 2012 / 131069 A1 describes short-chain fatty acid derivatives, such as propyl 3-hydroxypropionate and propyl propionate, for the treatment of immunogenic diseases, such as inflammatory diseases and viral infections (eg hepatitis).

[0006] PUCHALSKA PATRYCJA et al. (“Multi-dimensional Roles of Ketone Bodies in Fuel Metabolism, Signaling, and Therapeutics”, CELL METABOLISM, vol. 25, no. 2, 7 February 2017, pages 262-284) described the multiple therapeutic significances of ketone bodies, such as in multi-organ system inflammation and damage, heart failure, atherosclerosis, myocardial infarction, cancer, obesity, diabetes, NAFLD / NASH, neurological diseases, oxidative stress, etc.

[0007] Li et al. ("An exercise-inducible metabolite that suppresses feeding and obesity", NATURE, vol. 606, 15. June 2022, pages 785-790) N-lactylphenylalanine (Lac-Phe) discloses that exercise stimulates the production of N-lactyl-phenylalanine (Lac-Fhe), a blood-borne signaling metabolite that suppresses feeding and obesity. However, there is an urgent and currently unmet need for suitable lactate and ketone body preparations (e.g., BHB) to avoid excessive ion loads (e.g., sodium loads) and unpleasant taste. These factors severely limit the amount of metabolites that can be ingested, whether for daily intake as a food supplement or for use in a medically relevant setting.

[0008] Therefore, a compound or method of administering the compound that achieves the beneficial properties of lactate and BHB while minimizing harmful and unpleasant side effects is of particular importance. Summary of the invention

[0009] The present invention relates to compounds and compositions that can release lactic acid or lactic acid and BHB in vivo, referred to as the identification of lactic acid / ketone body esters. Therefore, the compounds of the present invention can be considered as prodrugs and / or nutraceuticals. Surprisingly, the inventors have also designed a new compound comprising pyruvic acid and BHB precursors, which are converted to lactic acid and BHB after ingestion and release. Therefore, without wishing to be bound by theory, the pyruvic acid-BHB compound (diPyKe) has the same effect as the lactic acid ketone body esters described herein.

[0010] Lactic acid / ketone body esters will have a dual function, limiting inflammation which is of general relevance, for example, to the treatment or amelioration of ageing-related diseases and they are high energy metabolic substrates. The latter effect is of relevance to medical uses, for example, for improving muscle function or output and / or limiting muscle atrophy in hospitalized patients or the general aging population. In addition, the esters may be relevant as food or nutritional supplements in, for example, endurance or high performance sports. In addition, these esters also have a range of further medical uses, such as reducing appetite / food intake in obese patients, which can also improve the conditions / symptoms of diabetes. Finally, these compounds are able to reduce free fatty acids and increase Lac-Phe, which is relevant to the treatment or reduction of obesity, diabetes and metabolic syndrome.

[0011] Therefore, one object of the present invention relates to providing compounds having the beneficial properties of lactic acid and BHB.

[0012] The object of the present invention is in particular to provide a compound that can solve the above problems without affecting its beneficial effects.

[0013] As used herein, unless otherwise indicated, the term "diLaKe" refers to a compound having the following general structure:

[0014]

[0015] As used herein, unless otherwise indicated, the term "LaKe" refers to a compound having the following general structure:

[0016]

[0017] wherein R1 is CH3 or OH, R2 is OH or H, and R3 is CH3 or H, preferably wherein R1, R2 and R3 are CH3, OH and H respectively, or R1, R2, R3 are OH, H and CH3 respectively.

[0018] As used herein, unless otherwise indicated, the term "DiLa" refers to a compound having the following general structure:

[0019]

[0020] wherein R1 is CH3 or OH, R2 is OH or H, and R3 is CH3 or H, preferably wherein R1, R2 and R3 are CH3, OH and H respectively, or R1, R2, R3 are OH, H and CH3 respectively.

[0021] As used herein, unless otherwise indicated, the term "KeLa" refers to a compound having the following general structure:

[0022]

[0023] wherein R1 is CH3 or OH, R2 is OH or H, and R3 is CH3 or H, preferably wherein R1, R2 and R3 are CH3, OH and H respectively, or R1, R2, R3 are OH, H and CH3 respectively.

[0024] As used herein, unless otherwise indicated, the term "diPyKe" refers to a compound having the following general structure:

[0025]

[0026] Preferably has the following stereochemical structure

[0027]

[0028] In a specific embodiment, diPyKe is (R)-butane-1,3-diylbis(2-oxopropionate).

[0029] Examples 1-2 show the synthesis of diLaKe compounds (Formula II).

[0030] Examples 3-5 describe the administration of diLaKe (Formula II) to rats via oral administration.

[0031] Examples 6-7 show the chemical synthesis of LaKe compounds XXII, XXIII, XXVIII and XXIX.

[0032] Example 8 shows the chemical synthesis of DiLa compounds XXVI and XXVII.

[0033] Examples 9-10 show that administration of LaKe (Formulas XXII and XXIII) by oral dosing resulted in increased serum concentrations of both BHB and lactate in rats compared to the control group.

[0034] Example 11 shows that oral administration of LaKe (Formula XXII and XXIII) resulted in a decrease in FFA concentration in rat serum compared to the control group.

[0035] Example 12 shows that administration of KeLa (Formulas XXIV and XXV) by oral dosing resulted in increased serum concentrations of both BHB and lactate in rats compared to the control group.

[0036] Example 13 shows that administration of DiLa (Formula XXVI and XXVII) by oral administration results in increased serum concentrations of lactic acid in rats compared to a control group.

[0037] Example 14 shows that oral administration of KeLa (Formulas XXIV and XXV) resulted in a decrease in FFA concentrations in rat serum compared to the control group.

[0038] Example 15 shows that oral administration of DiLa (Formula XXVI and XXVII) does not result in a decrease in FFA concentration in rat serum compared to the control group.

[0039] Examples 16-17 show the chemoenzymatic synthesis of LaKe (Formula XXII and XXIII) and DiLa (Formula XXVI and XXVII) compounds.

[0040] Example 18 shows the synthesis of R-(-)-β-hydroxybutyric acid ester (KeLa ester) (Formula XXIV and XXV).

[0041] Examples 19-20 show the chemical or chemoenzymatic synthesis of diLaKe.

[0042] Example 21 shows that oral administration of diLaKe results in increased serum concentrations of both BHB and lactate in rats.

[0043] Example 22 shows that oral administration of diLaKe results in a decrease in the concentration of FFA in rat serum.

[0044] Example 23 shows that oral administration of diLaKe (Formula II) results in increased serum concentrations of Lac-Phe.

[0045] Example 24 shows the chemical synthesis of diPyKe (Formula LXII).

[0046] Example 25 shows that oral administration of diPyKe results in increased serum concentrations of both BHB and lactate in rats.

[0047] Example 26 shows that oral administration of diPyKe (Formula LXII) results in increased serum concentrations of Lac-Phe.

[0048] Therefore, one aspect of the present invention relates to compounds of formula I

[0049]

[0050] or a pharmaceutically acceptable salt thereof.

[0051] Another aspect of the present invention relates to compounds of formula II or formula LXII

[0052] or

[0053]

[0054] or a pharmaceutically acceptable salt thereof.

[0055] Another aspect of the present invention relates to a composition comprising a compound described herein.

[0056] Another aspect of the present invention is to provide a pharmaceutical composition comprising a compound described herein or a composition described herein.

[0057] Another aspect of the present invention is to provide a compound as described herein, a composition as described herein or a pharmaceutical composition as described herein for use as a medicament.

[0058] Another aspect of the present invention is to provide a food ingredient comprising a compound as described herein or a composition as described herein.

[0059] Another aspect of the present invention is to provide a food product comprising the food ingredient described herein.

[0060] Another aspect of the invention is the use of a compound as described herein or a composition as described herein for non-therapeutic treatment.

[0061] BRIEF DESCRIPTION OF THE DRAWINGS

[0062] Figure 1

[0063] Figure 1 The results show that after oral administration of compounds XXII and XXIII (LaKe) or physiological saline solution (0.9% NaCl) to rats, the BHB ( Figure 1 A) and lactic acid ( Figure 1 B) Concentration (μM) over time.

[0064] Figure 2

[0065] Figure 2 The concentrations (μM) of non-esterified free fatty acids in the serum of rats after oral administration of compounds XXII and XXIII (LaKe) or a physiological saline solution (0.9% NaCl) as a control over time are shown.

[0066] Figure 3

[0067] Figure 3 The results show that after oral administration of compounds XXIV and XXV (KeLa) or physiological saline solution (0.9% NaCl) to rats, the BHB ( Figure 3 A) and lactic acid ( Figure 3 B) Concentration (μM) over time. The increase in lactate with KeLa ester was statistically significant starting at 90 min.

[0068] Figure 4

[0069] Figure 4 The results show that after oral administration of compounds XXVI and XXVII (DiLa) or physiological saline solution (0.9% NaCl) to rats, the BHB ( Figure 4 A) and lactic acid ( Figure 4 B) Concentration (μM) over time.

[0070] Figure 5

[0071] Figure 5 Shown are the concentrations (μM) of non-esterified free fatty acids in the serum of rats over time after oral administration of compounds XXIV and XXV (KeLa) or a physiological saline solution (0.9% NaCl) as a control.

[0072] Figure 6

[0073] Figure 6 The concentrations (μM) of non-esterified free fatty acids in the serum of rats after oral administration of compounds XXVI and XXVII (DiLa) or a physiological saline solution (0.9% NaCl) as a control over time are shown.

[0074] Figure 7

[0075] Figure 7 The results show that after oral administration of compound II (diLaKe) to rats, the BHB ( Figure 7 A) and lactic acid ( Figure 7 B) Concentration (μM) over time.

[0076] Figure 8

[0077] Figure 8 Shown is the concentration (μM) of non-esterified free fatty acids in rat serum over time after oral administration of Compound II (diLaKe) to rats.

[0078] Fig. 9

[0079] Fig. 9 Shown is the concentration (nM) of Lac-Phe in rat serum over time after oral administration of Compound II (diLaKe) to rats.

[0080] Fig.10

[0081] Fig.10 The results show that after oral administration of compound LXII (diPyKe) to rats, the BHB ( Fig.10 A) and lactic acid ( Fig.10 B) Concentration (μM) over time.

[0082] Fig.11

[0083] Fig.11 Shown is the concentration (nM) of Lac-Phe in rat serum over time after oral administration of compound LXII (diPyKe) to rats.

[0084] The present invention will be described in more detail below. DETAILED DESCRIPTION

[0085] definition

[0086] Before discussing the present invention in further detail, the following terms and conventions are first defined:

[0087] Ketone bodies

[0088] In this context, the term "ketone bodies" refers to water-soluble molecules or specific compounds BHB, acetoacetate and acetone that contain a ketone group. The compounds are produced by the liver from fatty acids.

[0089] Non-esterified free fatty acids (FFA)

[0090] In this context, the term "non-esterified free fatty acids" refers to metabolic fuels such as organic acids derived from the hydrolysis of endogenous triglycerides. Elevated levels of free fatty acids are associated with increased disease risk.

[0091] Oral administration

[0092] In the present context, the term "oral administration" refers to a route of administration in which a substance is administered through the oral cavity of a subject.

[0093] Subjects

[0094] The term "subject" includes humans of all ages, other primates (e.g., cynomolgus monkeys, rhesus monkeys); mammals in general, including commercially relevant mammals such as cattle, pigs, horses, sheep, goats, minks, ferrets, hamsters, cats and dogs, and birds. The preferred subject is a human.

[0095] The term "subject" also includes healthy subjects in the human population.

[0096] food

[0097] In the present context, the term "food" or "food ingredient" is to be understood as also covering "feed" and "feed ingredient". Thus, a food according to the present invention may refer to a food / feed suitable for human and / or animal consumption, and the term food ingredient according to the present invention may refer to a food / feed suitable for human and / or animal consumption.

[0098] Generally speaking, the term "feed" refers to animal consumption, while the term "food" refers to human or animal consumption. In the broad sense of the present invention, these terms can be used interchangeably.

[0099] Nutritional Supplements

[0100] In the present context, the term "nutraceutical" or "bio-nutraceutical" is to be understood as any substance that is a food or a part of a food and that can provide a medical or health benefit, including the prevention and treatment of disease.

[0101] A "nutraceutical" or "biogenic supplement" is a drug alternative that claims to have a physiological benefit. In the United States, "nutraceuticals" are largely unregulated because they fall under the authority of the Federal Food, Drug, and Cosmetic Act in the same category as FDA dietary supplements and food additives.

[0102] In a preferred embodiment of the present invention, the compound or composition of the present invention is a nutraceutical or a part of a nutraceutical.

[0103] In another preferred embodiment of the present invention, the compound or composition of the present invention is a medicament or a part of a medicament.

[0104] Lactic acid / ketone body esters

[0105] The present invention provides a group of novel lactic acid / ketone body esters and their synthesis for oral administration to animals and humans. These compounds provide the known beneficial properties of lactic acid and BHB while avoiding the toxic ion load associated with the use of the individual compounds.

[0106] Furthermore, compounds according to the present invention provide a unique solution for increasing serum concentrations of lactate (eg, (S)-lactate) and BHB (eg, (R)-BHB) in situations where delivery of relatively more lactate than BHB is desired.

[0107] Therefore, a first aspect of the present invention relates to compounds of formula I

[0108]

[0109] or a pharmaceutically acceptable salt thereof.

[0110] The compound of formula I contains an asymmetric C atom and can therefore exist in different isomers. In a preferred embodiment, the present invention relates to a compound of formula II

[0111]

[0112] or a pharmaceutically acceptable salt thereof.

[0113] In another embodiment, the present invention relates to compounds of formula III

[0114]

[0115] or a pharmaceutically acceptable salt thereof.

[0116] In another embodiment, the present invention relates to a compound of formula IV

[0117]

[0118] or a pharmaceutically acceptable salt thereof.

[0119] In another embodiment, the present invention relates to a compound of formula V

[0120]

[0121] or a pharmaceutically acceptable salt thereof.

[0122] In another embodiment, the present invention relates to a compound of formula VI

[0123]

[0124] or a pharmaceutically acceptable salt thereof.

[0125] In another embodiment, the present invention relates to a compound of formula VII

[0126]

[0127] or a pharmaceutically acceptable salt thereof.

[0128] In another embodiment, the present invention relates to a compound of formula VIII

[0129]

[0130] or a pharmaceutically acceptable salt thereof.

[0131] In another embodiment, the present invention relates to a compound of formula IX

[0132]

[0133] or a pharmaceutically acceptable salt thereof.

[0134] Finally, the present invention also provides another group of related novel pyruvic acid / keto body esters and their synthesis for oral administration to animals and humans. Without being bound by theory, these compounds provide the above-mentioned beneficial properties because the pyruvic acid released in the body is converted to lactic acid. In addition, the compound is an important intermediate in certain processes for producing diLaKe.

[0135] Therefore, another aspect of the present invention relates to compounds of formula LXI

[0136]

[0137] or a pharmaceutically acceptable salt thereof.

[0138] The compound of formula LXI contains an asymmetric C atom and can therefore exist as different isomers.

[0139] In a preferred embodiment, the present invention relates to compounds of formula LXII

[0140]

[0141] or a pharmaceutically acceptable salt thereof.

[0142] In another preferred embodiment, the present invention relates to compounds of formula LXIII

[0143]

[0144] or a pharmaceutically acceptable salt thereof.

[0145] Composition

[0146] The compounds of the invention may be used in compositions with other compounds, such as other lactic acid / ketone body esters.

[0147] Therefore, another aspect of the present invention relates to a composition comprising a compound of the present invention. In one embodiment, the present invention relates to a composition comprising a compound of formula II. In one embodiment, the present invention relates to a composition comprising a compound of formula LXII. In one embodiment, the present invention relates to a composition comprising a compound of formula II and a compound of formula LXII.

[0148] In another embodiment, the composition further comprises one or more compounds selected from the following group:

[0149] Compound of formula X

[0150]

[0151] Compound of formula XI

[0152]

[0153] and

[0154] Compound of formula XII

[0155]

[0156] or a pharmaceutically acceptable salt thereof,

[0157] wherein R1 is CH3 or OH, R2 is OH or H, and R3 is CH3 or H.

[0158] In another embodiment, the composition further comprises one or more compounds selected from the group consisting of:

[0159] Compound of formula X

[0160]

[0161] Compound of formula XI

[0162]

[0163] and

[0164] Compound of formula XII

[0165]

[0166] or a pharmaceutically acceptable salt thereof,

[0167] wherein R1, R2 and R3 are CH3, OH and H respectively, or R1, R2 and R3 are OH, H and CH3 respectively.

[0168] Formulas X, XI and XII have been tested in Examples 9-15.

[0169] In another embodiment, the composition further comprises one or more compounds selected from the following group:

[0170] Compound of formula XIII

[0171] Compound of formula XIV

[0172] Compound of formula XV

[0173] Compound of formula XVI

[0174] Compound of formula XVII

[0175] and the compound of formula XVIII

[0176] The compounds of Formula X, Formula XI and Formula XII may contain different asymmetric C atoms and thus may exist in different isomers.

[0177] In another embodiment, the composition further comprises one or more compounds selected from the group consisting of:

[0178] Compound of formula XIX

[0179]

[0180] Compound of formula XX

[0181]

[0182] Compound of formula XXI

[0183]

[0184] or a pharmaceutically acceptable salt thereof,

[0185] wherein R1, R2 and R3 are CH3, OH and H respectively, or R1, R2 and R3 are OH, H and CH3 respectively.

[0186] In another embodiment, the composition further comprises one or more compounds selected from the following group:

[0187] Compound of formula XXII

[0188]

[0189] Compound of formula XXIII

[0190] Compound of formula XXIV

[0191] Compound of formula XXV

[0192] Compound of formula XXVI

[0193] Compound of formula XXVII

[0194] Compound of formula XXVIII

[0195] and

[0196] Compound of formula XXIX

[0197]

[0198] or a pharmaceutically acceptable salt thereof.

[0199] In another embodiment, the composition further comprises a compound of formula XXXII,

[0200]

[0201] or a pharmaceutically acceptable salt thereof, wherein R1 is H, CH3, OH, CH2OH or CH(CH3)OH, R2 is H or OH, R3 is H or CH3, R4 and R5 are H, OH or CH3, and R6 is H or OH.

[0202] In one embodiment, R1 is CH(CH3)OH, R4 and R5 are H, and R6 is OH. In another embodiment, R1 is CH(CH3)OH, R4 and R5 are H, R6 is OH, and R2 is H. In one embodiment of the present invention, R1 is CH(CH3)OH. In another embodiment of the present invention, R2 is H. In another embodiment of the present invention, R3 is H. In one embodiment, R1 is CH2OH, R4 and R5 are H, and R6 is OH. In another embodiment, R1 is CH2OH, R4 and R5 are H, and R6 is OH. In another embodiment of the present invention, R4 is H. In another embodiment of the present invention, R5 is H. In another embodiment of the present invention, R6 is OH. In another embodiment, R1 is CH2OH. In another embodiment, R3 is CH3. In another embodiment, R1 is CH3. In another embodiment, R2 is OH. In a preferred embodiment, R1 is CH3, R2 is OH, R3 is H, R4 and R5 are H, and R6 is OH. In another embodiment, R1 is OH. In a preferred embodiment, R1 is OH, R2 is H, R3 is CH3, R4 and R5 are H, and R6 is OH. In one embodiment of the present invention, R4 is CH3. In another embodiment of the present invention, R4 is OH. In another embodiment of the present invention, R5 is CH3. In another embodiment of the present invention, R5 is OH. In another embodiment of the present invention, R6 is H.

[0203] In another embodiment, the composition further comprises a compound of formula XXXIII

[0204]

[0205] or a pharmaceutically acceptable salt thereof, wherein R1 is H, CH3, OH, CH2OH or CH(CH3)OH, R2 is H or OH, and R3 is H or CH3.

[0206] In another embodiment, the composition further comprises a compound of formula XXXIV

[0207]

[0208] or a pharmaceutically acceptable salt thereof, wherein R1 is H, CH3, OH, CH2OH or CH(CH3)OH, R2 is H or OH, and R3 is H or CH3.

[0209] In another embodiment, the composition further comprises a compound of formula XXXV

[0210]

[0211] or a pharmaceutically acceptable salt thereof, wherein R1 is H, CH3, OH, CH2OH or CH(CH3)OH, R2 is H or OH, and R3 is H or CH3, wherein R1 and R2 are different and are not H.

[0212] In another embodiment, the composition further comprises a compound of formula XXXVI

[0213]

[0214] or a pharmaceutically acceptable salt thereof, wherein R1 is H, CH3, OH, CH2OH or CH(CH3)OH, R2 is H or OH, and R3 is H or CH3, wherein R1 and R2 are different and are not H.

[0215] In another embodiment, the composition further comprises a compound of formula XXXVII

[0216] or a pharmaceutically acceptable salt thereof, wherein R1 is H, CH3, OH, CH2OH or CH(CH3)OH, R2 is H or OH, and R3 is CH3, wherein R1 and R2 are different and are not H.

[0217] In another embodiment, the composition further comprises a compound of formula XXXVIII

[0218]

[0219] or a pharmaceutically acceptable salt thereof, wherein R1 is H, CH3, OH, CH2OH or CH(CH3)OH, R2 is H or OH, and R3 is CH3, wherein R1 and R2 are different and are not H.

[0220] In another embodiment, the composition further comprises a compound of formula XXXIX

[0221]

[0222] or a pharmaceutically acceptable salt thereof, wherein R2 is H or OH, and R3 is H or CH3.

[0223] In another embodiment, the composition further comprises a compound of formula XXXX

[0224]

[0225] or a pharmaceutically acceptable salt thereof, wherein R2 is H or OH, and R3 is H or CH3.

[0226] In another embodiment, the composition further comprises a compound of formula XXXXI

[0227]

[0228] In another embodiment, the composition further comprises a compound of formula XXXXII

[0229] In another embodiment, the composition further comprises a compound of formula XXXXIII

[0230] In another embodiment, the composition further comprises a compound of formula XXXXIV

[0231] In another embodiment, the composition further comprises a compound of formula XXXXV

[0232] In another embodiment, the composition further comprises a compound of formula XXXXVI

[0233] In another embodiment, the composition further comprises a compound of formula XXXXVII

[0234] In another embodiment, the composition further comprises a compound of formula XXXXVIII

[0235]

[0236] In another embodiment, the composition further comprises a compound of formula XXXXIX

[0237]

[0238] In another embodiment, the composition further comprises a compound of formula L

[0239]

[0240] The compounds of the present invention and other compounds described above can be used in compositions containing different amounts of the compounds. The compounds of the present invention result in the production of two lactic acid molecules per BHB, therefore, combining the compounds with other compounds described herein will result in a variety of combinations having different lactic acid release characteristics relative to BHB.

[0241] In another embodiment, the composition comprises a compound of Formula I and a compound of Formula XXII.

[0242] In another embodiment, the composition comprises a compound of formula I and a compound of formula XXIII.

[0243] In another embodiment, the composition comprises a compound of formula I, a compound of formula XXII, and a compound of formula XXIII.

[0244] In another embodiment, the invention relates to a composition comprising a compound of formula I and a compound of formula XXII and / or XXIII, e.g., a ratio of formula I to formula XXII and / or XXIII in the range of 100:1 to 1:100, e.g., a ratio of 50:1 to 1:50, e.g., a ratio of 25:1 to 1:25, e.g., a ratio of 5:1 to 1:5 or 100:1 to 1:1, e.g., a ratio of 50:1 to 1:1, preferably a ratio of 25:1 to 1:1, more preferably a ratio of 25:1 to 5:1.

[0245] In another embodiment, the composition comprises a compound of formula II and a compound of formula XXII.

[0246] In another embodiment, the composition comprises a compound of formula II and a compound of formula XXIII.

[0247] In another embodiment, the composition comprises a compound of formula II, a compound of formula XXII, and a compound of formula XXIII.

[0248] In one embodiment, the invention relates to a composition comprising a compound of formula II and a compound of formula XXII and / or XXIII, for example, a ratio of formula II to formula XXII and / or XXIII in the range of 100:1 to 1:100, for example a ratio of 50:1 to 1:50, for example a ratio of 25:1 to 1:25, for example a ratio of 5:1 to 1:5 or 100:1 to 1:1, for example a ratio of 50:1 to 1:1, preferably a ratio of 25:1 to 1:1, more preferably a ratio of 25:1 to 5:1.

[0249] In another embodiment, the composition comprises a compound of Formula I and a compound of Formula XXIV.

[0250] In another embodiment, the composition comprises a compound of formula I and a compound of formula XXV. In another embodiment, the composition comprises a compound of formula I, a compound of formula XXIV, and a compound of formula XXV.

[0251] In another embodiment, the invention relates to a composition comprising a compound of formula I and a compound of formula XXIV and / or XXV, for example, a ratio of formula I to formula XXIV and / or XXV in the range of 100:1 to 1:100, for example, a ratio of 50:1 to 1:50, for example, a ratio of 25:1 to 1:25, for example, a ratio of 5:1 to 1:5 or 100:1 to 1:1, for example, a ratio of 50:1 to 1:1, preferably a ratio of 25:1 to 1:1, more preferably a ratio of 25:1 to 5:1.

[0252] In another embodiment, the composition comprises a compound of formula II and a compound of formula XXIV.

[0253] In another embodiment, the composition comprises a compound of Formula II and a compound of Formula XXV.

[0254] In another embodiment, the composition comprises a compound of formula II, a compound of formula XXIV, and a compound of formula XXV.

[0255] In one embodiment, the present invention relates to a composition comprising a compound of formula II and a compound of formula XXIV and / or XXV, for example, a ratio of formula II to formula XXIV and / or XXV in the range of 100:1 to 1:100, for example a ratio of 50:1 to 1:50, for example a ratio of 25:1 to 1:25, for example a ratio of 5:1 to 1:5 or 100:1 to 1:1, for example a ratio of 50:1 to 1:1, preferably a ratio of 25:1 to 1:1, more preferably a ratio of 25:1 to 5:1.

[0256] In another embodiment, the composition comprises a compound of Formula I and a compound of Formula XXVI.

[0257] In another embodiment, the composition comprises a compound of formula I and a compound of formula XXVII.

[0258] In another embodiment, the composition comprises a compound of formula I, a compound of formula XXVI, and a compound of formula XXVII.

[0259] In another embodiment, the invention relates to a composition comprising a compound of formula I and a compound of formula XXVI and / or XXVII, for example, a ratio of formula I to formula XXVI and / or XXVII in the range of 100:1 to 1:100, for example, a ratio of 50:1 to 1:50, for example, a ratio of 25:1 to 1:25, for example, a ratio of 5:1 to 1:5 or 100:1 to 1:1, for example, a ratio of 50:1 to 1:1, preferably a ratio of 25:1 to 1:1, more preferably a ratio of 25:1 to 5:1.

[0260] In another embodiment, the composition comprises a compound of formula II and a compound of formula XXVI.

[0261] In another embodiment, the composition comprises a compound of formula II and a compound of formula XXVII.

[0262] In another embodiment, the composition comprises a compound of formula II, a compound of formula XXVI, and a compound of formula XXVII.

[0263] In one embodiment, the present invention relates to a composition comprising a compound of formula II and a compound of formula XXVI and / or XXVII, for example, the ratio of formula II to formula XXVI and / or XXVII is in the range of 100:1 to 1:100, for example, the ratio is 50:1 to 1:50, for example, the ratio is 25:1 to 1:25, for example, the ratio is 5:1 to 1:5 or 100:1 to 1:1, for example, the ratio is 50:1 to 1:1, preferably the ratio is 25:1 to 1:1, more preferably the ratio is 25:1 to 5:1.

[0264] In another embodiment, the composition comprises a compound of formula LXI and a compound of formula XXII.

[0265] In another embodiment, the composition comprises a compound of formula LXI and a compound of formula XXIII.

[0266] In another embodiment, the composition comprises a compound of formula LXI, a compound of formula XXII, and a compound of formula XXIII.

[0267] In another embodiment, the present invention relates to a composition comprising a compound of formula LXI and a compound of formula XXII and / or XXIII, for example, a ratio of formula LXI to formula XXII and / or XXIII in the range of 100:1 to 1:100, for example a ratio of 50:1 to 1:50, for example a ratio of 25:1 to 1:25, for example a ratio of 5:1 to 1:5 or 100:1 to 1:1, for example a ratio of 50:1 to 1:1, preferably a ratio of 25:1 to 1:1, more preferably a ratio of 25:1 to 5:1.

[0268] In another embodiment, the composition comprises a compound of formula LXII and a compound of formula XXII.

[0269] In another embodiment, the composition comprises a compound of formula LXII and a compound of formula XXIII.

[0270] In another embodiment, the composition comprises a compound of formula LXII, a compound of formula XXII, and a compound of formula XXIII.

[0271] In one embodiment, the present invention relates to a composition comprising a compound of formula LXII and a compound of formula XXII and / or XXIII, for example, the ratio of formula LXII to formula XXII and / or XXIII is in the range of 100:1 to 1:100, for example, the ratio is 50:1 to 1:50, for example, the ratio is 25:1 to 1:25, for example, the ratio is 5:1 to 1:5 or 100:1 to 1:1, for example, the ratio is 50:1 to 1:1, preferably the ratio is 25:1 to 1:1, more preferably the ratio is 25:1 to 5:1.

[0272] In another embodiment, the composition comprises a compound of formula LXI and a compound of formula XXIV.

[0273] In another embodiment, the composition comprises a compound of formula LXI and a compound of formula XXV.

[0274] In another embodiment, the composition comprises a compound of formula LXI, a compound of formula XXIV, and a compound of formula XXV.

[0275] In another embodiment, the present invention relates to a composition comprising a compound of formula LXI and a compound of formula XXIV and / or XXV, for example, a ratio of formula LXI to formula XXIV and / or XXV in the range of 100:1 to 1:100, for example, a ratio of 50:1 to 1:50, for example, a ratio of 25:1 to 1:25, for example, a ratio of 5:1 to 1:5 or 100:1 to 1:1, for example, a ratio of 50:1 to 1:1, preferably a ratio of 25:1 to 1:1, more preferably a ratio of 25:1 to 5:1.

[0276] In another embodiment, the composition comprises a compound of formula LXII and a compound of formula XXIV.

[0277] In another embodiment, the composition comprises a compound of formula LXII and a compound of formula XXV.

[0278] In another embodiment, the composition comprises a compound of formula LXII, a compound of formula XXIV, and a compound of formula XXV.

[0279] In one embodiment, the present invention relates to a composition comprising a compound of formula LXII and a compound of formula XXIV and / or XXV, for example, the ratio of formula LXII to formula XXIV and / or XXV is in the range of 100:1 to 1:100, for example, the ratio is 50:1 to 1:50, for example, the ratio is 25:1 to 1:25, for example, the ratio is 5:1 to 1:5 or 100:1 to 1:1, for example, the ratio is 50:1 to 1:1, preferably the ratio is 25:1 to 1:1, more preferably the ratio is 25:1 to 5:1.

[0280] In another embodiment, the composition comprises a compound of formula LXI and a compound of formula XXVI.

[0281] In another embodiment, the composition comprises a compound of formula LXI and a compound of formula XXVII.

[0282] In another embodiment, the composition comprises a compound of formula LXI, a compound of formula XXVI, and a compound of formula XXVII.

[0283] In another embodiment, the present invention relates to a composition comprising a compound of formula LXI and a compound of formula XXVI and / or XXVII, for example, the ratio of formula LXI to formula XXVI and / or XXVII is in the range of 100:1 to 1:100, for example, a ratio of 50:1 to 1:50, for example, a ratio of 25:1 to 1:25, for example, a ratio of 5:1 to 1:5 or 100:1 to 1:1, for example, a ratio of 50:1 to 1:1, preferably a ratio of 25:1 to 1:1, more preferably a ratio of 25:1 to 5:1.

[0284] In another embodiment, the composition comprises a compound of formula LXII and a compound of formula XXVI.

[0285] In another embodiment, the composition comprises a compound of formula LXII and a compound of formula XXVII.

[0286] In another embodiment, the composition comprises a compound of formula LXII, a compound of formula XXVI, and a compound of formula XXVII.

[0287] In one embodiment, the invention relates to a composition comprising a compound of formula LXII and a compound of formula XXVI and / or XXVII, for example, the ratio of formula LXII to formula XXVI and / or XXVII is in the range of 100:1 to 1:100, for example, the ratio is 50:1 to 1:50, for example, the ratio is 25:1 to 1:25, for example, the ratio is 5:1 to 1:5 or 100:1 to 1:1, for example, the ratio is 50:1 to 1:1, preferably the ratio is 25:1 to 1:1, more preferably the ratio is 25:1 to 5:1.

[0288] In addition to the compounds disclosed in the present invention, the composition may further comprise other ketoesters or ketone body precursors.

[0289] Therefore, in one embodiment of the present invention, the composition further comprises one or more ketoesters or ketobody precursors, which are different from any compound of formula I to XXIX and XXXII to L, such as any compound of formula I to XXIX. In another embodiment of the present invention, the composition further comprises one or more ketoesters or ketobody precursors different from any compound of formula LXI to LXIII.

[0290] In another embodiment of the present invention, one or more ketoesters or ketobody precursors other than any of the compounds of formulae I to XXIX and XXXII to L, such as any of the compounds of formulae I to XXIX, are selected from 1,3-butanediol diacetate, 1,3-butanediol dihexanoate, 1,3-butanediol, medium chain triglycerides, medium chain fatty acids, 3-hydroxybutyl 3-hydroxybutyrate, and (3R)-hydroxybutyl (3R)-hydroxybutyrate. In another embodiment of the present invention, one or more ketoesters or ketobody precursors other than any of the compounds of formulae LXI to LXIII are selected from 1,3-butanediol diacetate, 1,3-butanediol dihexanoate, 1,3-butanediol, medium chain triglycerides, medium chain fatty acids, 3-hydroxybutyl 3-hydroxybutyrate, and (3R)-hydroxybutyl (3R)-hydroxybutyrate.

[0291] In another embodiment of the present invention, the composition described herein comprises a compound of formula II and at least one compound selected from 1,3-butanediol diacetate, 1,3-butanediol dihexanoate, 1,3-butanediol, medium chain triglycerides, medium chain fatty acids, 3-hydroxybutyl 3-hydroxybutyrate and (3R)-hydroxybutyl (3R)-hydroxybutyrate.

[0292] In another embodiment of the present invention, the composition described herein comprises a compound of formula LXII and at least one compound selected from 1,3-butanediol diacetate, 1,3-butanediol dihexanoate, 1,3-butanediol, medium chain triglycerides, medium chain fatty acids, 3-hydroxybutyl 3-hydroxybutyrate and (3R)-hydroxybutyl (3R)-hydroxybutyrate.

[0293] In another embodiment, the composition comprises a compound of formula II and (3R)-hydroxybutyl (3R)-hydroxybutyrate. In another embodiment, the composition comprises a compound of formula II and 1,3-butanediol dihexanoate. In another embodiment, the composition comprises a compound of formula II and 1,3-butanediol. In another embodiment, the composition comprises a compound of formula LXII and (3R)-hydroxybutyl (3R)-hydroxybutyrate. In another embodiment, the composition comprises a compound of formula LXII and 1,3-butanediol dihexanoate. In another embodiment, the composition comprises a compound of formula LXII and 1,3-butanediol dihexanoate. In another embodiment, the composition comprises a compound of formula LXII and 1,3-butanediol.

[0294] In another embodiment, the composition of the present invention further comprises a dietary and / or pharmaceutically acceptable carrier.

[0295] In another embodiment, the composition of the present invention further comprises a sugar carbohydrate.

[0296] use

[0297] BHB and lactic acid have been shown to improve or be useful in treating a number of different conditions. This is attributed to their anti-inflammatory effects, among other things, and their general role as high-energy substrates. Additionally, they will be associated with endurance or athletic performance, such as by improving muscle output / motor function.

[0298] Therefore, one aspect of the present invention relates to a pharmaceutical composition comprising a compound of the present invention or a composition of the present invention.

[0299] Thus, in one embodiment, the compounds described herein, the compositions described herein, or the pharmaceutical compositions described herein are for use as medicaments.

[0300] BHB and lactate have been associated with beneficial outcomes when used to treat inflammatory diseases, cancer, seizures, acute heart failure, resuscitation, acidosis, traumatic brain injury, acute pancreatitis, hepatitis, myocardial infarction, burns, sepsis, dengue fever, cognitive impairment, sarcopenia, atherosclerosis, neurodegeneration, oxidative stress, and wound healing. In addition, BHB and lactate have been associated with beneficial outcomes when used to treat chronic heart failure, cardiogenic shock, myocardial ischemia, pulmonary hypertension, systemic hypertension, organ transplant preparation, reperfusion injury after organ transplantation, and multiple sclerosis.

[0301] The compounds of formula XXII, XXIII, XXIV, XXV, XXVI and XXVII are capable of increasing circulating levels of ketone bodies BHB and / or lactate (see Examples 10, 12 and 13). The compounds of formula II and LXII are capable of increasing circulating levels of ketone bodies BHB and / or lactate (see Examples 21 and 25).

[0302] Therefore, in one embodiment of the present invention, the compounds described herein, the compositions described herein, or the pharmaceutical compositions described herein can be used to treat, prevent or alleviate inflammatory diseases, cancer, epileptic seizures, acute heart failure, resuscitation, acidosis, traumatic brain injury, acute pancreatitis, hepatitis, myocardial infarction, burns, sepsis, dengue fever, cognitive dysfunction, atherosclerosis, neurodegeneration, oxidative stress, Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, epilepsy, astrocytoma, glioblastoma and Huntington's disease, sarcopenia, muscle atrophy and wound healing dysfunction. In another embodiment of the present invention, the compounds described herein, the compositions described herein, or the pharmaceutical compositions described herein can be used to treat, prevent, or alleviate inflammatory diseases, cancer, epileptic seizures, acute and chronic heart failure, cardiogenic shock, resuscitation, acidosis, traumatic brain injury, acute pancreatitis, hepatitis, myocardial infarction, atherosclerosis, ischemic diseases, myocardial ischemia, pulmonary hypertension, systemic hypertension, organ transplant preparation, reperfusion injury after organ transplantation, burns, sepsis, dengue fever, cognitive dysfunction, atherosclerosis, neurodegeneration, oxidative stress, Alzheimer's disease, Parkinson's disease, multiple sclerosis, amyotrophic lateral sclerosis, epilepsy, astrocytoma, glioblastoma and Huntington's disease, sarcopenia, muscle atrophy, osteoporosis, wound healing dysfunction, damaged tissue, such as promoting damaged tissue repair, and infertility.

[0303] In another embodiment of the present invention, the compounds described herein, the compositions described herein, or the pharmaceutical compositions described herein can be used to treat, prevent, or reduce elevated plasma levels of free fatty acids in human or animal subjects; cognitive dysfunction, neurodegenerative diseases, such as Alzheimer's disease, Parkinson's disease, Huntington's disease, epilepsy; hypoxic states, such as angina pectoris, extreme physical exertion, intermittent claudication, hypoxia, stroke, and myocardial infarction; insulin resistance states, such as infection, stress, obesity, diabetes, metabolic syndrome, and heart failure; inflammatory states including infection and autoimmune diseases, as well as muscle damage, fatigue, and muscle fatigue. It is further disclosed that the compound reduces the plasma level of fatty acids and can be used, for example, to treat conditions caused, aggravated, or associated with elevated plasma levels of free fatty acids in human or animal subjects.

[0304] The compounds of formula XXII, XXIII, XXIV and XXV can reduce the level of free fatty acids (FFA) in the blood (see Examples 11 and 14). The compounds of formula II can reduce the level of free fatty acids (FFA) in the blood (see Example 22). Without wishing to be bound by theory, the compounds of formula LXII will have the same effect (Example 25).

[0305] FFA levels in the blood reflect ongoing lipolysis, and high levels of FFA induce insulin resistance; conversely, agents that reduce free fatty acid levels in the blood may be used therapeutically to increase insulin sensitivity in patients with type 2 diabetes and metabolic syndrome (6).

[0306] Therefore, in one embodiment of the present invention, the compounds, compositions or pharmaceutical compositions of the present invention can be used to treat, prevent or alleviate diseases caused by, aggravated by or associated with elevated plasma levels of free fatty acids in human or animal subjects.

[0307] N-lactylphenylalanine (Lac-Phe) is a lactic acid derived metabolite formed by the CDNP2 enzyme. Long-term administration of Lac-Phe to diet-induced obese mice can reduce fat and body weight and improve glucose homeostasis. Formula II compounds can increase the level of Lac-Phe in the blood (see Example 23). Without wishing to be bound by theory, Formula LXII compounds will have the same effect (Example 26).

[0308] Therefore, in one embodiment of the invention, the compounds, compositions or pharmaceutical compositions of the invention can be used to increase the level of appetite suppressant hormones, such as N-lactylphenylalanine (Lac-Phe). In one embodiment of the invention, the compounds, compositions or pharmaceutical compositions of the invention can be used to treat, prevent or alleviate a condition by increasing the plasma level of Lac-Phe in a human or animal subject. The method comprises administering a compound or composition of the invention to a subject.

[0309] Therefore, in a preferred embodiment of the present invention, these compounds can be used to control the level of free fatty acids in the blood of a subject.

[0310] In another embodiment of the invention, the compounds described herein, the compositions described herein, or the pharmaceutical compositions described herein may be used to treat, prevent, or alleviate viral infections, immunogenic diseases, non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), sarcopenia, muscle fatigue, angina, extreme physical exertion, intermittent claudication, hypoxia, stroke and myocardial infarction, stress, obesity, diabetes, metabolic syndrome, autoimmune diseases, muscle damage, or for improving motor function. In another embodiment of the invention, the compounds described herein, the compositions described herein, or the pharmaceutical compositions described herein may be used to treat, prevent, or alleviate viral infections, immunogenic diseases, non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), sarcopenia, muscle / lean body mass loss, muscle fatigue, angina, extreme physical exertion, intermittent claudication, hypoxia, stroke, CNS hemorrhage and myocardial infarction, stress, obesity, diabetes, metabolic syndrome, autoimmune diseases, muscle damage, or for improving motor function.

[0311] In one embodiment, the compounds, compositions or pharmaceutical compositions of the present invention can be used to treat, prevent and / or alleviate insulin resistance.

[0312] In another embodiment, the compounds described herein, the compositions described herein, or the pharmaceutical compositions described herein can be used to treat insulin resistance, preferably the compounds can be used to increase insulin sensitivity in a subject.

[0313] In another embodiment, the compounds described herein, the compositions described herein, or the pharmaceutical compositions described herein can be used in treatment to inhibit or reduce inflammation, atherosclerosis, neurodegeneration, oxidative stress, oncogenic angiogenesis, obesity, diabetes, and metabolic syndrome. In another embodiment, the compounds described herein, the compositions described herein, or the pharmaceutical compositions described herein can be used in treatment to inhibit or reduce inflammation, atherosclerosis, neurodegeneration, oxidative stress, oncogenic angiogenesis, obesity, diabetes, and metabolic syndrome.

[0314] Administration of the compounds described herein, compositions described herein, or pharmaceutical compositions described herein can be carried out in a variety of ways as described in the following non-limiting examples. Orally or nasally, i.e., by mouth or nose. By intradermal injection, the compound is delivered to the dermis of the skin between the epidermis and the subcutaneous tissue. Alternatively, the compound can be administered intravenously, which is administered directly into the bloodstream of the subject. In addition, intramuscular administration of the compound is injected into the muscle of the subject. In addition, the compound can be administered subcutaneously, which is under the skin, in the area between the muscle and skin of the subject. In addition, the compound can be administered intratracheally, which is administered directly into the trachea, and by transdermal administration, which is administered through the skin.

[0315] Any mode of administration can be used so long as it produces the desired effect of the compound.

[0316] Thus, in one embodiment, a compound described herein, a composition described herein, or a pharmaceutical composition described herein is administered to a subject orally or nasally.

[0317] In a preferred embodiment, a compound described herein, a composition described herein, or a pharmaceutical composition described herein is administered to a subject orally.

[0318] In another preferred embodiment, the compounds, compositions or pharmaceutical compositions of the present invention are administered orally.

[0319] A compound described herein, a composition described herein, or a pharmaceutical composition described herein can be administered at a dosage suitable to provide the desired effect in the subject receiving the compound.

[0320] In one embodiment, the compound described herein, the composition described herein, or the pharmaceutical composition described herein is administered at a dose of 0.05-1.5 g / kg, preferably 0.15-1.5 g / kg, more preferably 0.2-1.0 g / kg, even more preferably 0.2-0.5 g / kg.

[0321] In another embodiment, the compound, composition or pharmaceutical composition of the present invention is administered at a dose of 0.05-15 g / kg, preferably 0.15-10 g / kg, more preferably 0.2-5 g / kg, even more preferably 0.2-

[0322] 2.5 g / kg, even more preferably 0.2-1.5 g / kg.

[0323] In another embodiment, the compound, composition or pharmaceutical composition of the present invention is administered at a daily dose of 0.15-45 g / kg, preferably 0.45-30 g / kg, more preferably 0.6-15 g / kg or 0.6-4.5 g / kg, even more preferably 0.6-1.5 g / kg.

[0324] The "subject" described herein is assumed to receive the compound, including humans of all ages in need of the compound, other primates (e.g., cynomolgus monkeys, rhesus monkeys); general mammals, including commercially relevant mammals, such as cattle, pigs, horses, sheep, goats, minks, ferrets, hamsters, cats, dogs; and / or birds. The preferred subject is a human.

[0325] The term "subject" also includes healthy subjects in the human population.

[0326] Therefore, in one embodiment of the invention, the subject is selected from the following groups; humans of all ages, other primates (e.g., cynomolgus monkeys, rhesus monkeys); general mammals, including commercially relevant mammals such as cattle, pigs, horses, sheep, goats, minks, ferrets, hamsters, cats and dogs, and birds.

[0327] In a preferred embodiment, the subject is a human.

[0328] In one embodiment, the subject is an elderly subject, such as a person over the age of 40, such as over the age of 50, such as over the age of 60, such as over the age of 70, or such as over the age of 80. In one embodiment, the subject has a high body mass index (BMI), such as a BMI> 30. Thus, without being bound by theory, in some embodiments, a high BMI can be used to distinguish between therapeutic and non-therapeutic treatments.

[0329] In one embodiment, the compounds of the invention are administered as part of a pharmaceutical composition.

[0330] The compounds and compositions of the present invention may be in different forms, such as dry powder, aqueous solution, or gel.

[0331] In one embodiment of the invention, the compound described herein, the composition described herein, or the pharmaceutical composition described herein is an aqueous solution, a gel, or a powder, preferably in an aqueous solution.

[0332] Food ingredients and food products

[0333] The compounds or compositions of the present invention may be included in food ingredients.

[0334] Thus, one aspect of the present invention relates to a food ingredient comprising a compound or composition of the present invention.

[0335] Furthermore, the food ingredient of the present invention may be part of a food product. Thus, one aspect of the present invention relates to a food product comprising the food ingredient of the present invention.

[0336] In one embodiment of the present invention, the food is selected from the group consisting of nutraceuticals, food supplements, dietary supplements, feeds, bars, candy bars, protein bars, powders, gels, beverages, alcoholic beverages, yogurts, chewing gums, dairy products, sports drinks, candy products, ice cream, capsules, tablets, granules and pouches.

[0337] Non-therapeutic use

[0338] The compounds or compositions of the present invention may be used in non-therapeutic treatments.

[0339] Thus, one aspect of the present invention relates to the use of a compound or composition of the present invention for reducing circulating free fatty acids in the plasma of a subject in the non-therapeutic treatment of muscle damage fatigue or to improve motor function.

[0340] In one embodiment of the present invention, the compound or composition is used for non-therapeutic treatment of myocardial fatigue, skeletal muscle fatigue and / or improvement of motor function, or for improving alertness or improving cognitive function of a subject.

[0341] In one embodiment of the present invention, the compound or composition is used for non-therapeutic treatment of myocardial fatigue, skeletal muscle fatigue and / or improvement of motor function, or for improving alertness or improving cognitive function of a subject, or preventing neurodegeneration in a subject.

[0342] In one embodiment of the present invention, the compound or composition can promote the mental health of a subject.

[0343] In one embodiment, the composition for reducing circulating free fatty acids in the plasma of a subject in the non-therapeutic treatment of muscle damage or fatigue does not comprise DiLa.

[0344] Therefore, one aspect of the present invention relates to the use of a compound or composition of the present invention for suppressing appetite, treating obesity, promoting weight loss, preventing, reducing and / or treating sarcopenia, maintaining a healthy weight or reducing the fat to lean muscle ratio.

[0345] One aspect also relates to the use of a compound or composition of the present invention for non-therapeutic increase of Lac-Phe serum concentration.

[0346] In one embodiment of the invention, the compound or composition can increase the lifespan of a subject.

[0347] In one embodiment of the present invention, the compound or composition is used to promote wound healing and tissue repair after injury in a subject.

[0348] In one embodiment of the invention, the compound or composition is used to promote bone health in a subject, such as for non-therapeutic treatment of osteoporosis, such as non-therapeutic increase of bone mass.

[0349] method

[0350] The compounds can be produced by chemoenzymatic synthesis as described in Example 1 or Example 20 or chemical synthesis as shown in Example 2 or Example 19.

[0351] Therefore, in one aspect, the present invention relates to a method for producing a compound of the present invention, the method comprising:

[0352] a) providing a compound of formula XXX;

[0353]

[0354] b) reacting the compound of step a) with ethyl lactate, such as (S)-(-)-ethyl lactate, in the presence of lipase; and

[0355] c) providing a compound as described herein.

[0356] In one embodiment, the present invention is a method for producing a compound of formula II, the method comprising:

[0357] a) providing a compound of formula XXXI;

[0358]

[0359] b) reacting the compound of step a) with ethyl lactate, such as (S)-(-)-ethyl lactate, in the presence of lipase; and

[0360] c) providing a compound of formula II.

[0361] In one embodiment, the method further comprises step d) purifying the provided compound, for example by filtration, distillation and / or flash column chromatography (FCC).

[0362] In another embodiment, the lipase is immobilized on a solid support, such as beads.

[0363] Lipases catalyze the hydrolysis of triacylglycerols into glycerol and free fatty acids. However, lipases may also be used for other functions, such as in the above-mentioned methods.

[0364] Candida antarctica lipase B (CALB) is considered a major enzyme in biotechnology due to its broad substrate specificity, high activity, and high enantioselectivity. It also has the ability to work in both aqueous and non-aqueous reaction environments. CALB can be used for transesterification, kinetic resolution, and polymerization reactions.

[0365] Therefore, in another embodiment, the lipase is selected from lipase B, lipase B immobilized on Immobead150, and lipase B andida antarctica. 435, CALB, CALB lipase immobilized on a hydrophobic support, LipozymeTL IM and RM, 1,3-specific lipase.

[0366] In a preferred embodiment, the lipase is lipase B, more preferably CALB or Novozym 435.

[0368] In the Examples section (Example 1), Novozym 435 was used. In the Examples section (Examples 16 and 17), the lipase B Candida Antarctica immobilized on Immobead 150 was used.

[0369] As described in Example 20, the inventors designed an alternative chemoenzymatic approach that first provides diPyKe via a chemical synthesis step.

[0370] Therefore, in another aspect, the present invention relates to a method for producing a compound of the present invention, the method comprising:

[0371] a) providing 1,3-butanediol, such as (R)-1,3-butanediol and pyruvic acid;

[0372] b) reacting the compound of step a) in the presence of a catalytic acid, such as p-toluenesulfonic acid (pTsOH); and

[0373] c) obtaining diPyKe as described herein.

[0374] In one embodiment, step b is carried out in a suitable solvent, such as anhydrous toluene.

[0375] To further provide diLaKe, the method may include an additional hydrogenation step to reduce the ketone of diPyKe to an alcohol. In one embodiment, the method includes step d) of reacting diPyKe from step c in the presence of a dehydrogenase. In one embodiment, the method includes step d) of reacting diPyKe from step c in the presence of a stoichiometric reducing agent or another hydrogenation method, such as by catalytic hydrogenation.

[0376] Thus, this aspect may be defined as a method of producing a compound of the invention, the method comprising:

[0377] a) providing 1,3-butanediol, such as (R)-1,3-butanediol and pyruvic acid;

[0378] b) reacting the compound of step a) in the presence of a catalytic acid, such as p-toluenesulfonic acid (pTsOH);

[0379] c) obtaining the diPyKe of the present invention;

[0380] d) reacting the compound of step c) in the presence of a reducing agent such as a dehydrogenase, for example in the presence of baker's yeast;

[0381] e) obtaining diLaKe as described herein.

[0382] In one embodiment, the dehydrogenase is selected from alcohol dehydrogenase, lactate dehydrogenase, pyruvate dehydrogenase and glutamate dehydrogenase. The dehydrogenase may be from yeast. Preferably, the dehydrogenase is purified so that it consists essentially of purified enzymes. When a purified dehydrogenase is used, it may also be necessary to include a suitable coenzyme, such as NADH or NADPH, to allow the reaction to function, and may include means for re-fermenting the coenzyme.

[0383] The source of the dehydrogenases may also be provided by their presence in yeast cells such as baker's yeast. Thus, in one embodiment, the dehydrogenase is baker's yeast, such as YSC2 from Sigma-Aldrich.

[0384] In one embodiment, the method further comprises the step of purifying the provided compound, for example by filtration, distillation and / or flash column chromatography (FCC).

[0385] In another aspect, the invention relates to a step of producing said compound, wherein

[0386]

[0387] First, alkyl (R7) lactate, such as methyl lactate, ethyl lactate or propyl lactate is converted into formula A compound. Hydroxyl is protected with a protecting group (P) known to those skilled in the art, for example by using TBSCl, imidazoles and DMF. P can be any protecting group well known in the art. Those skilled in the art will know suitable protecting groups for hydroxyl, examples of which can be based on trialkylsilyl derivatives, such as trimethylsilyl (TMS), triethylsilyl (TES), tert-butyldimethylsilyl (TBS), tert-butyldiphenylsilyl (TBDPS) and triisopropylsilyl (TIPS). Protecting groups can be based on removable groups under hydrogenation conditions, such as benzyl (Bn), p-methoxybenzyl (p-OMe-Mn). Protecting groups can be based on removable groups under acidic conditions, such as methoxymethyl (MOM), benzyloxymethyl (BOM).

[0388] In the second step, the compound of formula A is converted into a compound of formula B by hydrolyzing the ester group using, for example, an alkaline solution obtained by dissolving LiOH, NaOH, KOH, CsOH, K3PO4, Cs2CO3 in a suitable solvent. Suitable solvents for this reaction step may be tetrahydrofuran (THF) or ethanol. However, other solvents may also be used, such as alcohols such as 2-methyl THF (2-Me-THF), water, ethanol or isopropanol, acetonitrile, 1,4-dioxane, dimethyl sulfoxide (DMSO), N,N-dimethylformamide (DMF), diethylene glycol dimethyl ether (bis(2-methoxyethyl) ether) and a mixture of solvents. After hydrolysis, the mixture is neutralized with an acid, and product B is isolated using standard extraction procedures known to those skilled in the art.

[0389] Next, the compound of formula C is prepared by reacting 1,3-butanediol with the compound of formula B in the presence of a coupling agent or dehydrating agent such as O-(benzotriazol-1-yl)-N,N,N',N'-tetramethyluronium tetrafluoroborate (TBTU) to form the compound of formula C. Any coupling agent or dehydrating agent known in the art may be used, such as N,N'-dicyclohexylcarbodiimide (DCC), N,N'-diisopropylcarbodiimide (DIC), N-ethyl-N'-(3-dimethylaminopropyl)carbodiimide (EDC), N,N'-dibutylsuccinimidyl carbonate (DSC), (1-cyano-2-ethoxy-2-oxoethylenaminooxy)dimethylamino-morpholinium-carbonium hexafluorophosphate (COMU), O-(benzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HBTU), tetramethylfluorouronium hexafluorophosphate (TFFH). This may also involve forming an activated species, such as a mixed anhydride, from formula B by reacting formula B with a chloroformate or an acyl halide. The reaction can be carried out in a solvent such as dimethylformamide (DMF) or other solvents known to those skilled in the art, and may further contain an organic compound such as N,N-diisopropylethylamine (DIPEA), triethylamine (TEA), 4-dimethylaminopyridine (DMAP) or other bases known to those skilled in the art.

[0390] Finally, the protecting group (P) is removed from the compound of formula C by methods known in the art for various protecting groups defined herein, such as TBS can be removed under acidic conditions, such as by KHSO4, HCl, or by reacting with a fluoride source, such as tetrabutylammonium fluoride (TBAF), hydrogen fluoride, hydrogen fluoride pyridine complex, triethylamine trifluoride. Suitable solvents for this reaction step can be tetrahydrofuran (THF), methanol and water or a mixture thereof. However, other solvents such as acetonitrile, 2-Me-THF, toluene, ethyl acetate, dichloromethane can also be used alone or as a mixture. Another example is to remove benzyl by adding palladium carbon (Pd / C) and a hydrogen source (such as H2). Suitable solvents for this reaction step can be ethanol and water or a mixture thereof. However, other solvents such as methanol, tetrahydrofuran (THF), 2-Me-THF, ethyl acetate (EtOAc), toluene, DMF, acetonitrile can also be used alone or as a mixture. Alternative metal catalysts can be used, including their fixed forms, such as platinum, rhodium or nickel. Furthermore, an alternative method involves transfer hydrogenation using 2-propanol and formate as hydrogen sources.

[0391] In addition, the final product (diLake) can be purified by methods well known in the art, such as by removing MeOH or EtOH under reduced pressure, and extracting with EtOAc after adding water, collecting the organic layer and drying it with Na2SO4, filtering and concentrating, followed by flash column chromatography on silica gel, crystallization, precipitation or distillation.

[0392] Other aspects

[0393] For example, US201123766A1 discloses a number of physical conditions or diseases that can be treated with ketone compounds.

[0394] One aspect of the invention relates to a method of treating a disease caused by, exacerbated by or associated with elevated plasma levels of free fatty acids in a human or animal subject, the method comprising administering to the subject a compound, composition or pharmaceutical composition of the invention.

[0395] Another aspect of the present invention relates to a method of treating a condition associated with weight loss or weight gain, the method comprising administering to a subject in need thereof a compound, composition or pharmaceutical composition of the present invention.

[0396] Another aspect of the invention relates to a method of suppressing appetite, treating obesity, promoting weight loss, maintaining healthy weight, or reducing fat to lean muscle ratio, the method comprising administering to a subject in need thereof a compound, composition, or pharmaceutical composition of the invention.

[0397] Another aspect of the present invention relates to a method for preventing or treating a condition selected from cognitive dysfunction, neurodegenerative diseases or disorders, muscle damage, fatigue and muscle fatigue, comprising administering a compound, composition or pharmaceutical composition of the present invention to a subject in need thereof.

[0398] One aspect relates to a method of treating a patient suffering from a disorder selected from diabetes, hyperthyroidism, metabolic syndrome X, or treating an elderly patient, the method comprising administering thereto a compound, composition or pharmaceutical composition of the present invention.

[0399] Another aspect relates to a method of treating, preventing, or reducing the effects of neurodegeneration, free radical toxicity, hypoxic conditions, or hyperglycemia, the method comprising administering a compound, composition, or pharmaceutical composition of the invention to a subject in need thereof. In one embodiment, neurodegeneration is caused by aging, trauma, hypoxia, or a neurodegenerative disease or disorder.

[0400] One aspect also relates to a method for preventing or treating a neurodegenerative disease or disorder selected from Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, epilepsy, astrocytoma, glioblastoma and Huntington's disease, the method comprising administering to a subject in need thereof a compound, composition or pharmaceutical composition of the present invention.

[0401] Another aspect relates to a method of promoting alertness or improving cognitive function in a subject, comprising administering to the subject a compound, composition or pharmaceutical composition of the present invention.

[0402] One aspect also relates to the use of a compound or composition of the invention for reducing free fatty acids circulating in the plasma of a subject in the non-therapeutic treatment of muscle damage or fatigue.

[0403] One aspect also relates to the use of a compound or composition of the present invention for non-therapeutic increase of Lac-Phe serum concentration.

[0404] Another aspect relates to the use of the compounds or compositions of the present invention, wherein the compounds or compositions are used for the non-therapeutic treatment of myocardial fatigue or skeletal muscle fatigue. Another aspect relates to the use of the compounds or compositions of the present invention, wherein the compounds or compositions are used for the non-therapeutic treatment of heart failure.

[0405] The present invention relates to a lactate / ketone body ester for protecting vital organ function and combating inflammation and cancer growth. In particular, the present invention relates to a lactate / ketone body ester having the beneficial properties of lactate and beta-hydroxybutyrate (BHB) on vital organ function, inflammation and cancer growth, but without the harmful sodium load produced by the administration of lactate and beta-hydroxybutyrate.

[0406] The present invention relates to lactate / ketone body esters for protecting vital organ function and combating inflammation, cancer growth, sarcopenia and atherosclerosis. In particular, the present invention relates to lactate / ketone body esters having the beneficial properties of lactate or lactate and ketone bodies such as beta-hydroxybutyrate (BHB) on vital organ function, inflammation and cancer growth, but without the harmful ion loads such as calcium, potassium, magnesium and / or sodium loads following administration of lactate and beta-hydroxybutyrate.

[0407] Invented Projects

[0408] 1. A compound of formula I,

[0409]

[0410] or a pharmaceutically acceptable salt thereof;

[0411] or a compound of formula LXII

[0412]

[0413] 2. A compound according to item 1 having the stereochemistry of formula II

[0414]

[0415] or a pharmaceutically acceptable salt thereof.

[0416] 3. A composition comprising any of the above compounds.

[0417] 4. The composition according to item 3, which comprises a compound of formula II.

[0418] 5. The composition according to any one of items 3-4, further comprising one or more compounds selected from the following groups:

[0419] The compound of formula X,

[0420]

[0421] The compound of formula XI,

[0422]

[0423] and

[0424] Compound of formula XII

[0425]

[0426] or a pharmaceutically acceptable salt thereof,

[0427] wherein R1, R2 and R3 are CH3, OH and H respectively, or R1, R2 and R3 are OH, H and CH3 respectively.

[0428] 6. The composition according to any one of items 3 to 5, further comprising one or more compounds selected from the following groups:

[0429] Compound of formula XXII

[0430]

[0431] Compound of formula XXIII

[0432]

[0433] Compound of formula XXIV

[0434]

[0435] Compound of formula XXV

[0436]

[0437] Compound of formula XXVI

[0438]

[0439] Compound of formula XXVII

[0440]

[0441] Compound of formula XXVIII

[0442]

[0443] and

[0444] Compound of formula XXIX

[0445]

[0446] or a pharmaceutically acceptable salt thereof.

[0447] 7. The composition according to any one of items 3 to 6, comprising a compound of formula II, a compound of formula XXII and / or a compound of formula XXIII.

[0448] 8. A composition according to any one of items 3 to 7, comprising a compound of formula II and at least one compound selected from 1,3-butanediol diacetate, 1,3-butanediol dihexanoate, 1,3-butanediol, medium-chain triglycerides, medium-chain fatty acids, 3-hydroxybutyl 3-hydroxybutyrate and (3R)-hydroxybutyl (3R)-hydroxybutyrate.

[0449] 9. A pharmaceutical composition comprising the compound described in any one of items 1 to 2 or the composition described in any one of items 3 to 8.

[0450] 10. The compound according to any one of items 1 to 2, the composition according to any one of items 3 to 8 or the pharmaceutical composition according to item 9 for use as a medicament.

[0451] 11. The compound according to any one of items 1-2, the composition according to any one of items 3-8 or the pharmaceutical composition according to item 9, for use

[0452] - Treat, prevent or alleviate inflammatory diseases, cancer, epileptic seizures, acute heart failure, resuscitation, acidosis, traumatic brain injury, acute pancreatitis, hepatitis, myocardial infarction, burns, sepsis, dengue fever, cognitive dysfunction, atherosclerosis, neurodegeneration, oxidative stress, Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, epilepsy, astrocytoma, glioblastoma and Huntington's disease, sarcopenia, muscle atrophy, wound healing dysfunction;

[0453] - Treating, preventing or alleviating conditions caused by, aggravated by or associated with elevated plasma free fatty acid levels in human or animal subjects;

[0454] - Treat, prevent or alleviate viral infections, immunogenic disorders, non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), sarcopenia, muscle fatigue, angina pectoris, extreme physical exertion, intermittent claudication, hypoxia, stroke and myocardial infarction, stress, obesity, diabetes, metabolic syndrome, autoimmune diseases, muscle damage or for improving motor function; and / or

[0455] -Treat, prevent and / or reduce insulin resistance.

[0456] 12. A food ingredient comprising the compound according to any one of items 1 to 2 or the composition according to any one of items 3 to 8.

[0457] 13. A food product comprising the food ingredient according to item 12.

[0458] 14. The food product according to item 13, wherein the food is selected from the group consisting of nutritional products, food supplements, dietary supplements, feeds, bars, candy bars, protein bars, powders, gels, beverages, alcoholic beverages, yogurts, chewing gums, dairy products, sports drinks, candy products, ice cream, capsules, tablets, granules and sachets.

[0459] 15. Use of a compound according to any one of items 1 to 2 or a composition according to any one of items 3 to 8,

[0460] - Reduction of circulating free fatty acids in the plasma of subjects in non-therapeutic treatments for muscle damage, fatigue or to improve motor function;

[0461] - Suppress appetite, treat obesity, promote weight loss, prevent, reduce and / or treat sarcopenia, maintain a healthy weight or reduce the ratio of fat to lean muscle; and / or

[0462] - for non-therapeutic treatment of myocardial fatigue, skeletal muscle fatigue, and / or improvement of motor function, or for promoting alertness or improving cognitive function in a subject;

[0463] It should be noted that embodiments and features described in the context of one aspect of the invention may also be applicable to other aspects of the invention.

[0464] All patent and non-patent references cited in this application are incorporated herein by reference in their entirety.

[0465] The invention will now be described in more detail in the following non-limiting examples.

[0466] Example

[0467] Example 1 - Chemoenzymatic Synthesis of diLaKe

[0468] Purpose

[0469] Provides a chemoenzymatic synthesis of diLaKe.

[0470] Materials, Methods & Results

[0471]

[0472] The procedure for the chemoenzymatic synthesis of diLaKe is as follows:

[0473] (R)-1,3-Butanediol (0.9 g), (S)-ethyl lactate (11.8 g) and Novozym 435 (0.5 g) were placed in a sealed vial and gently vortexed at 40°C for 40 h. At this time, GC analysis showed 90% conversion of (R)-1,3-butanediol, and 20% diLaKe. After removing the excess (S)-ethyl lactate by distillation, analytically pure diLaKe was obtained by flash column chromatography on silica gel.

[0474] CalB can be used in place of Novozym 435 in the synthesis. 10% diLaKe was produced as determined by GC analysis after 40 h.

[0475] 1 H NMR

[0476] diLaKe:(400MHz,CDCl3)δ H 5.11(m,1H),4.24(m,4H),1.97(m,2H),1.41(m,6H),1.30(m,3H)

[0477] Using NMR spectroscopy, it was confirmed that diLaKe was produced as described above.

[0478] in conclusion

[0479] diLaKe can be produced by chemoenzymatic synthesis.

[0480] Example 2 - Chemical Synthesis of diLaKe

[0481] Purpose

[0482] Provide chemical synthesis of diLaKe

[0483] Materials and methods

[0484]

[0485]

[0486] The procedure for the synthesis of diLaKe is as follows:

[0487] To a solution of (S)-ethyl lactate (14.4 mL, 127 mmol, 1.0 eq.) in anhydrous DMF (250 mL) was added TBCl (28.7 g, 191 mmol, 1.5 eq.), followed by imidazole (30.3 g, 445 mmol, 3.5 eq.). The mixture was stirred at room temperature for 3 hours, then diluted with a saturated aqueous NaCl solution and extracted with EtOAc. The organic layers were combined and washed with a 5% aqueous hydrochloric acid solution and a saturated aqueous NaCl solution. The organic layer was then dried over Na2SO4, filtered and concentrated to give (S)-2-(tert-butyldimethylsilyloxy)propanoate A (29.5 g, quantitative). By 1 H NMR analysis confirmed the formation of the product. The product was directly carried forward to the next step without further purification.

[0488] R f

[0489] 0.41 (pentane / EtOAc 98:2, KMnO4).

[0490] 1 H NMR

[0491] (400MHz,CDCl3)δH 4.34-4.26(q,J=6.74Hz,1H),4.24-4.11(m,2H),1.41-1.36(d,J=6.75Hz,3H), 1.30-1.24(t,J=7.16Hz,3H),0.91-0.87(s,9H),0.11-0.04(d,J=11.99Hz,6H).

[0492] To a solution of (S)-2-(tert-butyldimethylsilyloxy)propanoate A (29.5 g, 127 mmol, 1.0 eq.) in THF (400 mL) was added cooled LiOH aqueous solution (508 mL, 0.5 M) at 0°C. The reaction mixture was stirred at room temperature for 5 hours, then concentrated to half of the original volume and extracted with Et2O. The organic extracts were combined and then extracted with saturated NaHCO3 aqueous solution. The aqueous layers were combined and acidified with 1M KHSO4 aqueous solution until pH ≈ 3-4 was reached. The aqueous solution was then thoroughly extracted with Et2O, the organic layers were combined, dried over Na2SO4 and concentrated to give (S)-2-(tert-butyldimethylsilyloxy)propanoic acid B (22.3 g, 86%) as an oil. By 1 H NMR analysis confirmed the formation of the product ( 1 H NMR values ​​were consistent with reported values) (10).

[0493] 1 H NMR

[0494] (400MHz,CDCl3)δ H 4.38-4.33(q,J=6.88Hz,1H),1.46-1.45(d,J=6.80Hz,3H),0.94-0.90(m,9H),0.16-0.12(s,6H).

[0495] (S)-2-(tert-butyldimethylsilyloxy)propionic acid B (2.1 eq.), TBTU (3.0 eq.) and DIPEA (4.2 eq.) were dissolved in anhydrous DMF and the mixture was stirred at room temperature for 1 hour. A solution of (R)-(-)-1,3-butanediol (1.0 eq.) in anhydrous DMF was added and the reaction mixture was stirred overnight at ambient temperature. The reaction mixture was diluted with CH2Cl2 and the resulting mixture was washed sequentially with 1M aqueous HCl solution, aqueous NaHCO3 solution and water. The organic layers were then combined, dried over Na2SO4, filtered and concentrated to give the crude product. Silica gel flash column chromatography (or alternative purification method) was performed to give the pure product (C).

[0496] To a stirred solution of C (1.0 eq.) in THF:H2O:MeOH (2.5:2.5:1) was added KHSO4 (0.25 eq.) and the mixture was stirred at room temperature. MeOH was removed under reduced pressure, water was added, and then extracted with EtOAc. The organic layer was collected, dried over Na2SO4, filtered and concentrated. Silica gel flash column chromatography (or alternative purification method) was performed to obtain the pure product diLaKe.

[0497] result

[0498] Using NMR spectroscopy, it was confirmed that diLaKe was produced as described above.

[0499] in conclusion

[0500] diLaKe can be produced by chemical synthesis.

[0501] Example 3 - Animal Experiment Design

[0502] Preparation of drugs

[0503] The diLaKe ester (Formula II) was diluted in saline solution (0.9% NaCl) and administered as a single bolus of 2 mL per oral dose. The dose of 4500 mg / kg was based on a previous test series. Placebo animals received a single bolus of 2 mL of saline solution (0.9% NaCl).

[0504] Animal setup:

[0505] Male Sprague-Dawley rats were housed at 23°C for acclimatization with a 12-h light-dark cycle and unlimited access to food and water. All animal handling was in accordance with the Danish national guidelines and the Guide for the Care and Use of Laboratory Animals. 1 ,All experiments were carried out in accordance with Danish law (Act No. 1306 of 23 November 2007).

[0506] Rats were randomly selected to receive diLaKe or placebo.

[0507] In the induction chamber, 8% sevoflurane ( Rats were anesthetized with 1x100 μl / mL flow rate (2.0 L / min) using AbbVIE A / S, Copenhagen, Denmark.

[0508] After anesthesia was achieved, the rats were intubated and connected to a mechanical ventilator (Ugo Basile 7025 rodent ventilator, Comerio, Varese, Italy) with a flow rate of 1.0 L / min containing 3.5% sevoflurane. The body temperature was maintained at a constant 37°C ± 1°C using a temperature probe (UNO, Zevenaar, Holland). A PTFE-coated flexible orogastric tube (Fuchigami, Japan) was placed and the rats were allowed to stabilize for 15 minutes.

[0509] After stabilization, baseline blood samples were collected from the tail vein of rats and then diLaKe or placebo was administered.

[0510] Following the baseline blood sample, animals were administered a dose of diLaKe solution or placebo via an orogastric tube. Blood samples (200 uL each) were collected in microcontainers (sarstedt–

[0511] 20.1280.100), allowed to clot for 30 minutes, and then centrifuged at 1500G for 20 minutes at 4°C. Serum was collected and stored at -80°C for further analysis.

[0512] Example 4 - Quantification of BHB and lactate in rat serum - diLaKe (Formula II)

[0513] The concentrations of BHB and lactate were determined in blood samples isolated from rats according to Example 3. Rat serum isolated every 15 minutes was analyzed for 2 hours. Quantification was performed on LC-MS / MS using an isotopically labeled internal standard (7).

[0514] in conclusion

[0515] Here, it is shown that treatment with diLaKe (Formula II) results in increased serum concentrations of both BHB and lactate in rats following oral administration compared to the control group (saline 0.9%), similar to the increase in serum concentrations following treatment with LaKe in Example 10.

[0516] Example 5-Effect of dilake (Formula II) on free fatty acids

[0517] The concentration of free fatty acids (FFA) in blood samples isolated from rats according to Example 3 was determined.

[0518] Blood levels of FFA reflect ongoing lipolysis, and high levels of FFA induce insulin resistance; conversely, agents that reduce blood levels of free fatty acids may be used therapeutically to increase insulin sensitivity in people with type 2 diabetes and metabolic syndrome.

[0519] The concentration of non-esterified free fatty acids (NEFA) was measured using the NEFA-HR (2) kit (Wako, Chemicals Gmbh). The absorbance was measured by spectroscopy (PHERAstar FS, BMG LABTECH, Ortenberg, Germany).

[0520] The concentration of free fatty acids (mM) in the serum of rats treated with diLaKe was expected to be reduced compared to rats treated with the control group (saline 0.9%).

[0521] in conclusion

[0522] Here, it is shown that oral administration of diLaKe (Formula II) results in a decrease in FFA concentrations in rat serum compared to the control group, similar to the decrease in concentrations following treatment with LaKe in Example 11.

[0523] Example 6 - Chemical Synthesis of LaKe Compounds 3 and 4 (Formula XXII and XXIII)

[0524] The LaKe compound was prepared according to the following procedure.

[0525] Synthesis of compounds XXII and XXIII:

[0526]

[0527] (S)-2-((tert-Butyldimethylsilyl)oxy)propanoic acid (1):

[0528]

[0529] To a solution of (S)-2-hydroxypropionic acid ethyl ester (14.4 mL, 127 mmol, 1.0 eq.) in anhydrous DMF (250 mL) was added TBCl (28.7 g, 191 mmol, 1.5 eq.), followed by imidazole (30.3 g, 445 mmol, 3.5 eq.). The mixture was stirred at room temperature for 3 hours, then diluted with saturated aqueous NaCl solution and extracted with EtOAc. The organic layers were combined and washed with 5% aqueous HCl solution and saturated aqueous NaCl solution. The organic layer was then dried over Na2SO4, filtered and concentrated to give (S)-2-(tert-butyldimethylsilyloxy)propionic acid ethyl ester (29.5 g, quantitative). By 1 H NMR analysis confirmed the formation of the product. The product was directly carried into the next step without further purification.

[0530] R f

[0531] 0.41 (pentane / EtOAc 98:2, KMnO4).

[0532] 1 H NMR

[0533] (400MHz,CDCl3)δ H 4.34-4.26(q,J=6.74Hz,1H),4.24-4.11(m,2H),1.41-1.36(d,J=6.75Hz,3H), 1.30-1.24(t,J=7.16Hz,3H),0.91-0.87(s,9H),0.11-0.04(d,J=11.99Hz,6H).

[0534] To a solution of (S)-2-(tert-butyldimethylsilyloxy)propanoate (29.5 g, 127 mmol, 1.0 eq.) in THF (400 mL) was added cooled LiOH aqueous solution (508 mL, 0.5 M) at 0°C. The reaction mixture was stirred at room temperature for 5 hours, then concentrated to half of the original volume and extracted with Et2O. The organic extracts were combined and then extracted with saturated aqueous NaHCO3. The aqueous layers were combined and acidified with 1 M aqueous KHSO4 to reach pH ≈ 3-4. The aqueous solution was then thoroughly extracted with Et2O, and the organic layers were combined, dried over Na2SO4 and concentrated to give (S)-2-(tert-butyldimethylsilyloxy)propanoic acid (22.3 g, 86%) as an oil. 1 H NMR analysis confirmed the formation of the product ( 1 H NMR values ​​were consistent with reported values) (10).

[0535] 1 H NMR

[0536] (400MHz,CDCl3)δ H 4.38-4.33(q,J=6.88Hz,1H),1.46-1.45(d,J=6.80Hz,3H),0.94-0.90(m,9H),0.16-0.12(s,6H).

[0537] (R)-3-Hydroxybutyl (S)-2-((tert-)butyldimethylsilyl)oxy)propionate (2):

[0538]

[0539] (S)-2-(tert-butyldimethylsilyloxy)propionic acid (8.42 g, 41.2 mmol, 1.0 eq.), TBTU (19.9 g, 61.8 mmol, 1.5 eq.) and DIPEA (15.1 mL, 86.5 mmol, 2.1 eq.) were dissolved in anhydrous DMF (150 mL), and the mixture was stirred at room temperature for 1 hour. A solution of (R)-(-)-1,3-butanediol (3.68 mL, 41.2 mmol, 1.0 eq.) in anhydrous DMF (10 mL) was then added, and the reaction mixture was stirred overnight at ambient temperature. The reaction mixture was diluted with CH2Cl2, and the resulting mixture was washed with 1M HCl aqueous solution, NaHCO3 aqueous solution and water in sequence. The organic layers were then combined, dried over Na2SO4, filtered and concentrated to give a crude product. FCC (pentane / EtOAc=95:5 to 85:15) was performed to give the pure product (2) (5601 mg, 49%).

[0540] R f

[0541] 0.38 (pentane / EtOAc 8:2, KMnO4).

[0542] 1 H NMR

[0543] (400MHz,CDCl3)δ H 4.42-4.30(m,2H),4.21-4.16(m,1H),3.93-3.84(m,1H),2.00-1.97(d,J=4.27Hz,1H),1.85-1.69(m,2H),1 .42-1.38(d,J=6.79Hz,3H),1.24-1.21(d,J=6.27Hz,3H),0.91-0.89(s,9H),0.11-0.07(d,J=10.17Hz,6H).

[0544] 13 C NMR

[0545] (101MHz,CDCl3)δ C 174.6,68.6,65.1,62.3,38.2,25.9,23.6,21.5,18.4,-4.8,-5.1.

[0546] IR(neat)

[0547] υ max / cm -1 3440,2930,2858,1737,1463,1373,1256,1140.

[0548] HRMS

[0549] [M+Na] + =299.1649; found:299.1644.

[0550] [α] 24.0 D

[0551] -34.6°(C=10mg / mL,CHCl3).

[0552] (R)-3-Hydroxybutyl (S)-2-hydroxypropionate (3) and (R)-4-hydroxybutan-2-yl (S)-2-hydroxypropionate (4):

[0553]

[0554] To a stirred solution of TBDMS ether (5612 mg, 20.3 mmol, 1.0 eq.) in THF:H2O:MeOH (2.5:2.5:1, 100 mL) was added KHSO4 (691 mg, 5.08 mmol, 0.25 eq.), and the mixture was stirred at room temperature overnight. MeOH was removed under reduced pressure, water was added, and then extracted with EtOAc. The organic layer was collected, dried over Na2SO4, filtered and concentrated. The crude product was purified by FCC (pentane / EtOAc=1:1) to obtain the desired product (2100 mg, 68%, r.10:1).

[0555] R f

[0556] 0.19 (pentane / EtOAc 1:1, CAM).

[0557] 1 H NMR

[0558] 3:(400MHz,CDCl3)δ H 4.46-4.37(m,1H),4.32-4.23(m,2H),3.95-3.85(m,1H),2.81-2.79(d,J=5.28Hz, 1H),1.87-1.71(m,3H),1.44-1.39(d,J=6.97Hz,3H),1.27-1.22(d,J=6.53Hz,3H).

[0559] 4:(400MHz,CDCl3)δ H5.25-5.17(m,1H),4.46-4.37(m,1H),3.74-3.57(m,2H),1.87-1.71(m,2H),1.44-1.39(d,J=6.97Hz,3H),1.33-1.30(d,J=6.22Hz,3H).

[0560] 13 C NMR

[0561] 3:(101MHz,CDCl3)δ C 176.1,66.9,64.9,63.0,38.0,23.8,20.6.

[0562] IR(neat)

[0563] υ max / cm -1 3380,2971,1732,1457,1375,1274,1209,1130,1049.

[0564] HRMS

[0565] [M+Na] + =185.0784; found:185.0783.

[0566] Example 7 - Chemical Synthesis of LaKe Compounds 10 and 11 (Formula XXVIII and XXIX)

[0567] The LaKe compound was prepared according to the following procedure.

[0568] Synthesis of compounds XXVIII and XXIX:

[0569]

[0570] (R)-2-((tert-Butyldimethylsilyl)oxy)propanoic acid (9):

[0571]

[0572] To a solution of (R)-ethyl 2-hydroxypropanoate (6.74 mL, 59.3 mmol, 1.0 eq.) in anhydrous DMF (100 mL) was added TBSCl (13.4 g, 89 mmol, 1.5 eq.), followed by imidazole (14.1 g, 208 mmol, 3.5 eq.). The mixture was stirred at room temperature for 3 hours, then diluted with saturated aqueous NaCl solution and extracted with EtOAc. The organic layers were combined, washed with 5% aqueous HCl solution and saturated aqueous NaCl solution. The organic layer was then dried over Na2SO4, filtered and concentrated to give ethyl (R)-2-(tert-butyldimethylsilyloxy)propanoate (13.7 g, quantitative). 1 H NMR analysis confirmed product formation. The product was directly carried into the next step without further purification.

[0573] R f

[0574] 0.41 (pentane / EtOAc 98:2, KMnO4).

[0575] 1 H NMR

[0576] (400MHz,CDCl3)δ H 4.34-4.26(q,J=6.74Hz,1H),4.24-4.11(m,2H),1.41-1.36(d,J=6.75Hz,3H), 1.30-1.24(t,J=7.16Hz,3H),0.91-0.87(s,9H),0.11-0.04(d,J=11.99Hz,6H).

[0577] To a solution of (R)-2-(tert-butyldimethylsilyloxy)propanoate (13.7 g, 59.3 mmol, 1.0 eq.) in THF (400 mL) was added cooled LiOH aqueous solution (400 mL, 0.3 M) at 0°C. The reaction mixture was stirred at room temperature for 5 hours, then concentrated to half of its original volume and extracted with Et2O. The organic extracts were combined and then extracted with saturated aqueous NaHCO3. The aqueous layers were combined and acidified with 1 M aqueous KHSO4 to pH ≈ 3-4. The aqueous solution was then thoroughly extracted with Et2O, and the organic layers were combined, dried over Na2SO4 and concentrated to give (R)-2-(tert-butyldimethylsilyloxy)propanoic acid (9085 mg, 75%) as an oil. 1 H NMR analysis confirmed the formation of the product ( 1 H NMR values ​​are consistent with reported values) 1 .

[0578] 1 H NMR

[0579] (400MHz,CDCl3)δ H 4.38-4.33(q,J=6.88Hz,1H),1.46-1.45(d,J=6.80Hz,3H),0.94-0.90(m,9H),0.16-0.12(s,6H).

[0580] (R)-3-Hydroxybutyl (R)-2-hydroxypropionate (10) and (R)-4-hydroxybutan-2-yl (R)-2-hydroxypropionate (11):

[0581]

[0582] (R)-2-((tert-butyldimethylsilyl)oxy)propionic acid (8.99 g, 44.0 mmol, 1.0 eq.), TBTU (21.3 g, 66.0 mmol, 1.5 eq.) and DIPEA (16.1 mL, 92.4 mmol, 2.1 eq.) were dissolved in anhydrous DMF (175 mL) and the mixture was stirred at room temperature for 1 hour. A solution of (R)-(-)-1,3-butanediol (3.93 mL, 44.0 mmol, 1.0 eq.) in anhydrous DMF (10 mL) was then added and the reaction mixture was stirred overnight at ambient temperature. The reaction mixture was diluted with CH2Cl2 and the resulting mixture was washed with 1M HCl aqueous solution, NaHCO3 aqueous solution and water in sequence. The organic layers were then combined, dried over Na2SO4, filtered and concentrated to give a crude product. FCC (pentane / EtOAc = 95:5 to 85:15) gave the pure product (5576 mg, 46%).

[0583] R f

[0584] 0.38 (pentane / EtOAc 8:2, KMnO4).

[0585] 1 H NMR

[0586] (400MHz,CDCl3)δ H4.44-4.36(m,1H),4.36-4.29(q,J=6.58Hz,1H),4.20-4.13(m,1H),3.93-3.82(m,1H),2.01-1.96(d,J=4.44Hz,1H),1.86- 1.67(m,2H),1.42-1.38(d,J=6.69Hz,3H),1.25-1.20(d,J=6.21Hz,3H),0.93-0.87(s,9H),0.11-0.07(d,J=10.35Hz,6H).

[0587] IR(neat)

[0588] υ max / cm -1 3449,2958,2930,2900,2858,1736,1253,1138.

[0589] HRMS

[0590] [M+Na] + =299.1649; found:299.1658.

[0591] [α] 24.0 D

[0592] +17.1°(C=8.5mg / mL,CHCl3).

[0593] To a stirred solution of TBDMS ether (5576 mg, 20.2 mmol, 1.0 eq.) in THF:H2O:MeOH (2.5:2.5:1, 100 mL) was added KHSO4 (668 mg, 5.05 mmol, 0.25 eq.), and the mixture was stirred at room temperature overnight. MeOH was removed under reduced pressure, water was added, and then extracted with EtOAc. The organic layer was collected, dried over Na2SO4, filtered and concentrated. The crude product was purified by FCC (pentane / EtOAc=1:1) to obtain the desired product (2110 mg, 64%, r.10:1).

[0594] R f

[0595] 0.19 (pentane / EtOAc 1:1, CAM).

[0596] 1 H NMR

[0597] 10:(400MHz,CDCl3)δ H4.49-4.38(m,1H),4.33-4.20(m,2H),3.95-3.83(m,1H),2.82-2.74(d,J=4.96Hz, 1H),1.98-1.61(m,3H),1.44-1.39(d,J=6.92Hz,3H),1.27-1.22(d,J=6.22Hz,3H).

[0598] 11:(400MHz,CDCl3)δ H 5.25-5.17(m,1H),4.49-4.38(m,1H),3.71-3.64(m,1H),3.63-3.54(m,1H),1 .98-1.61(m,3H),1.44-1.39(d,J=6.92Hz,3H),1.34-1.20(d,J=6.21Hz,3H).

[0599] 13 C NMR

[0600] 10:(101MHz,CDCl3)δ C 176.1,66.9,64.9,63.0,37.9,23.7,20.5.

[0601] IR(neat)

[0602] υ max / cm -1 3377,2971,2901,1732,1376,1209,1128,1079,1049.

[0603] HRMS

[0604] [M+Na] + =185.0784; found:185.0790.

[0605] Example 8 - Synthesis of DiLa compounds 7 and 8 (Formula XXVI and XXVII)

[0606] Synthesis of compounds XXVI and XXVII:

[0607]

[0608] (S)-2-Hydroxypropyl (S)-2-((tert-butyldimethylsilyl)oxy)propionate (5) and (S)-1-hydroxypropan-2-yl (S)-2-((tert-butyldimethylsilyl)oxy)propionate (6):

[0609]

[0610] (S)-2-(tert-butyldimethylsilyloxy)propionic acid (5.80 g, 28.4 mmol, 1.0 eq.), TBTU (13.7 g, 42.6 mmol, 1.5 eq.) and DIPEA (10.4 mL, 59.6 mmol, 2.1 eq.) were dissolved in anhydrous DMF (80 mL) and the mixture was stirred at room temperature for 1 hour. A solution of (S)-(-)-1,2-propylene glycol (2.08 mL, 28.4 mmol, 1.0 eq.) in anhydrous DMF (10 mL) was then added and the reaction mixture was stirred overnight at ambient temperature. The reaction mixture was diluted with CH2Cl2 and the resulting mixture was washed with 1M HCl aqueous solution, NaHCO3 aqueous solution and water in sequence. The organic layers were then combined, dried over Na2SO4, filtered and concentrated to give a crude product. FCC (pentane / EtOAc=85:15) afforded 5 and 6 (2850 mg, 38%; r.10:1).

[0611] R f

[0612] 0.14 (pentane / EtOAc 9:1, KMnO4).

[0613] 1 H NMR

[0614] 5:(400MHz,CDCl3)δ H 4.40-4.32(q,J=6.79Hz,1H),4.16-4.10(dd,J=10.79Hz,2.76Hz,1H),4.08-4.01(m,1H),4.01-3.94(dd,J=10.77Hz,7.13Hz,1H),2.1 4-2.09(d,J=3.47Hz,1H),1.44-1.39(d,J=6.72Hz,3H),1.23-1.18(d,J=6.26Hz,3H),0.92-0.88(s,9H),0.12-0.05(d,J=9.51Hz,6H).

[0615] 6:(400MHz,CDCl3)δ H5.05-4.96(m,1H),4.34-4.29(q,J=6.69Hz,1H),3.71-3.58(m,2H),1.96-1.91(t,J=6.05Hz,1H),1.41 -1.38(d,J=6.72Hz,3H),1.26-1.23(d,J=6.56Hz,3H),0.92-0.88(s,9H),0.12-0.05(d,J=9.51Hz,6H).

[0616] 13 C NMR

[0617] 5:(101MHz,CDCl3)δ C 174.3,70.0,68.5,66.2,25.8,21.4,19.2,18.4,-4.8,-5.1.

[0618] IR(neat)

[0619] υ max / cm -1 3508,2953,2931,2887,2857,1739,1472,1254,1203,1141,1061.

[0620] HRMS

[0621] [M+Na] + =285.1493; found:285.1497.

[0622] (S)-2-Hydroxypropyl (S)-2-hydroxypropionate (7) and (S)-1-hydroxypropan-2-yl (S)-2-hydroxypropionate (8):

[0623]

[0624] To a stirred solution of TBDMS ether (2834 mg, 10.8 mmol, 1.0 eq.) in THF:H2O:MeOH (2.5:2.5:1, 50 mL) was added KHSO4 (368 mg, 2.70 mmol, 0.25 eq.), and the mixture was stirred at room temperature overnight. MeOH was removed under reduced pressure, water was added, and then extracted with EtOAc. The organic layer was collected, dried over Na2SO4, filtered and concentrated. The crude product was purified by FCC (pentane / EtOAc=1:1 to 4:6) to obtain the desired product (880 mg, 55%, r.3:1).

[0625] R f

[0626] 0.12 (Pentane / EtOAc 1:1, KMnO4).

[0627] 1 H NMR

[0628] 7: (400 MHz, CDCl3) δ H 4.37 - 4.29 (q, J=6.62 Hz, 1H), 4.18 - 4.12 (dd, J=10.70 Hz, 2.54 Hz, 1H), 4.08 - 4.00 (m, 1H), 4.00 - 3.94 (dd, J=10.71 Hz, 7.28 Hz, 1H), 3.90 - 3.83 (s, 1H), 3.38 - 3.30 (s, 1H), 1.44 - 1.40 (d, J=6.99 Hz, 3H), 1.21 - 1.17 (d, J=6.14 Hz, 3H).

[0629] 8: (400 MHz, CDCl3) δ H 5.06 - 4.97 (m, 1H), 4.30 - 4.24 (q, J=6.81 Hz, 1H), 3.86 - 3.80 (m, 1H), 3.70 - 3.63 (d, J=11.81 Hz, 2H), 3.63 - 3.53 (m, 1H), 3.28 - 3.19 (s, 1H), 1.42 1.38 (d, J=6.72 Hz, 3H), 1.25 - 1.21 (d, J=6.43 Hz, 3H).

[0630] 13 C NMR

[0631] 7: (101 MHz, CDCl3) δ C 175.3, 70.2, 67.2, 65.8, 20.4, 19.0.

[0632] 8: (101 MHz, CDCl3) δ C 175.3, 73.1, 67.4, 65.3, 20.4, 16.1.

[0633] IR(neat)

[0634] υ max / cm -1 3391, 2984, 1734, 1455, 1374, 1213, 1130, 1054.

[0635] HRMS

[0636] [M + Na] +=171.0628; found:171.0629.

[0637] Example 9 - Design of animal experiment

[0638] Preparation of drugs

[0639] LaKe esters (Formula XXII and XXIII) were diluted with saline solution (0.9% NaCl) and administered as a single bolus of 2 mL per oral dose. The dose of 4500 mg / kg was based on a previous test series (data not shown). Placebo animals received a single bolus of 2 mL of saline solution (0.9% NaCl).

[0640] Animal settings :

[0641] Male Sprague-Dawley rats (300-350 g, Taconic, Ry, Denmark) were housed in a 3-hour environment at 23°C with a 12-h light-dark cycle and unlimited access to food and water. All animal handling was in accordance with the Danish national guidelines and the Guide for the Care and Use of Laboratory Animals. 1 ,All experiments were carried out in accordance with Danish law (Act No. 1306 of 23 November 2007).

[0642] Rats were randomly selected to receive LaKe (n=8) or placebo (n=8).

[0643] In the induction chamber, 8% sevoflurane ( Rats were anesthetized with 1x100 μl / mL flow rate (2.0 L / min) using AbbVIE A / S, Copenhagen, Denmark.

[0644] After anesthesia was achieved, the rats were intubated and connected to a mechanical ventilator (Ugo Basile 7025 rodent ventilator, Comerio, Varese, Italy) with a flow rate of 1.0 L / min containing 3.5% sevoflurane. The body temperature was maintained at a constant 37°C ± 1°C using a temperature probe (UNO, Zevenaar, Holland). A PTFE-coated flexible orogastric tube (Fuchigami, Japan) was placed and the rats were allowed to stabilize for 15 minutes.

[0645] After stabilization, baseline blood samples were collected from the tail vein of rats and then LaKe or placebo was administered.

[0646] After the baseline blood sample, the animals were administered a dose of LaKe solution or placebo via an orogastric tube. Blood samples (200 uL each) were collected in microcontainers (sarstedt–

[0647] 20.1280.100), allowed to clot for 30 minutes, and then centrifuged at 1500G for 20 minutes at 4°C. Serum was collected and stored at -80°C for further analysis.

[0648] Statistical analysis

[0649] Based on our own experience and reports from other research groups, a sample size of n = 8 was considered sufficient to determine treatment effects. All results are expressed as mean ± SD unless otherwise stated. Differences in concentrations were analyzed using two-way ANOVA with Bonferroni post hoc correction. All analyses were performed using GraphPad Prism 8.2.0 (Graph Pad Software, CA, USA). P < 0.05 was considered statistically significant.

[0650] Example 10 - Quantification of BHB and lactic acid in rat serum - LaKe (Formula XXII and XXIII)

[0651] The concentrations of BHB and lactate were determined in blood samples isolated from rats according to Example 4. Rat serum isolated every 15 minutes was analyzed for 2 hours. Quantification was performed on LC-MS / MS using an isotopically labeled internal standard (7).

[0652] Figure 1 A: BHB concentration (μM) in rat sera isolated from two groups of rats, one treated with LaKe and the other with control treatment.

[0653] Figure 1 B: Lactate concentration (μM) in rat sera isolated from two groups of rats, one treated with LaKe and the other with control treatment.

[0654] in conclusion

[0655] Here, we show that treatment with LaKe (Formulae XXII and XXIII) results in increased serum concentrations of both BHB and lactate after oral administration in rats compared to the control group (saline 0.9%).

[0656] Example 11 - Effect of LaKe Esters (Formula XXII and XXIII) on Free Fatty Acids

[0657] The concentration of free fatty acids (FFA) was determined in blood samples isolated from rats according to Example 4. The level of FFA in the blood reflects ongoing lipolysis, and high levels of FFA induce insulin resistance; conversely, agents that reduce the level of free fatty acids in the blood can be used therapeutically to increase insulin sensitivity in people with type 2 diabetes and metabolic syndrome.

[0658] The concentration of non-esterified free fatty acids (NEFA) was measured using the NEFA-HR (2) kit (Wako, Chemicals Gmbh). The absorbance was measured by spectroscopy (PHERAstar FS, BMG LABTECH, Ortenberg, Germany).

[0659] Figure 2 : The concentration of free fatty acids (mM) in the serum of rats treated with LaKe was significantly reduced compared with rats treated with control (normal saline 0.9%).

[0660] in conclusion

[0661] Here, we show that oral administration of LaKe (Formula XXII and XXIII) resulted in a decrease in FFA concentrations in rat serum compared to the control group.

[0662] Example 12 - Quantification of BHB and lactate in rat serum - KeLa (Formulas XXIV and XXV)

[0663] The concentrations of BHB and lactate were determined in blood samples isolated from rats according to Example 4, except that KeLa (Formulas XXIV and XXV) was used. Rat serum isolated every 15 minutes was analyzed for 2 hours. Quantification was performed on LC-MS / MS using an isotope-labeled internal standard (7).

[0664] Figure 3 A: BHB concentration (μM) in rat sera isolated from two groups of rats, one treated with KeLa and the other with control treatment.

[0665] Figure 3 B: Lactate concentration (μM) in rat sera isolated from two groups of rats, one treated with KeLa and the other with control treatment. The increase in lactate with KeLa ester was statistically significant from 90 minutes and beyond.

[0666] in conclusion

[0667] Here, we show that treatment with KeLa (Formulae XXIV and XXV) results in increased serum concentrations of both BHB and lactate in rats following oral administration compared to the control group (saline 0.9%).

[0668] Example 13 - Quantification of BHB and lactic acid in rat serum - DiLa (Formula XXVI and XXVII)

[0669] The concentrations of BHB and lactate were determined in blood samples isolated from rats according to Example 4, except that DiLa (Formulas XXVI and XXVII) was used. Rat serum isolated every 15 minutes was analyzed for 2 hours. Quantification was performed on LC-MS / MS using an isotope-labeled internal standard (7).

[0670] Figure 4 A: BHB concentration (μM) in rat sera isolated from two groups of rats, one treated with DiLa and one treated with control.

[0671] Figure 4 B: Lactate concentration (μM) in rat sera isolated from two groups of rats, one treated with DiLa and one treated with control.

[0672] in conclusion

[0673] Here, we show that treatment with DiLa (Formulae XXVI and XXVII) results in increased serum lactate concentrations in rats after oral administration compared to the control group (saline 0.9%).

[0674] DiLa esters do not appear to release BHB directly (only lactate), but the data suggest that BHB mobilization was inhibited during the experimental period relative to the control group. This is consistent with the apparent inhibition of ketone formation (ketogenesis) in the liver, as FFA levels, if any, which are precursors to ketone bodies, are elevated after DiLa ester administration.

[0675] Example 14 - Effect of KeLa Esters (Formula XXIV and XXV) on Free Fatty Acids

[0676] The concentration of free fatty acids (FFA) in blood samples isolated from rats according to Example 4 was determined, except that KeLa (Formulas XXIV and XXV) was used. The level of FFA in the blood reflects ongoing lipolysis, and high levels of FFA induce insulin resistance; conversely, agents that reduce the level of free fatty acids in the blood can be used therapeutically to increase insulin sensitivity in people with type 2 diabetes and metabolic syndrome.

[0677] The concentration of non-esterified free fatty acids (NEFA) was measured using the NEFA-HR (2) kit (Wako, Chemicals Gmbh). The absorbance was measured by spectroscopy (PHERAstar FS, BMG LABTECH, Ortenberg, Germany).

[0678] Figure 5 : The concentration of free fatty acids (μM) in the serum of rats treated with Kela was significantly reduced compared to rats treated with control (saline 0.9%).

[0679] in conclusion

[0680] Here, we show that oral administration of Kela (Formulas XXIV and XXV) resulted in a decrease in FFA concentrations in rat serum compared to the control group.

[0681] Example 15 - Effect of DiLa Esters (Formula XXVI and XXVII) on Free Fatty Acids

[0682] The concentration of free fatty acids (FFA) in blood samples isolated from rats according to Example 4 was determined, except that DiLa (Formulas XXVI and XXVII) was used. The level of FFA in the blood reflects ongoing lipolysis, and high levels of FFA induce insulin resistance; conversely, agents that reduce the level of free fatty acids in the blood can be used therapeutically to increase insulin sensitivity in people with type 2 diabetes and metabolic syndrome.

[0683] The concentration of non-esterified free fatty acids (NEFA) was measured using the NEFA-HR (2) kit (Wako, Chemicals Gmbh). The absorbance was measured by spectroscopy (PHERAstar FS, BMG LABTECH, Ortenberg, Germany).

[0684] Figure 6 : The concentration of free fatty acids (μM) in the serum of rats treated with DiLa was not reduced compared with rats treated with control (saline 0.9%).

[0685] in conclusion

[0686] Here, we show that oral administration of DiLa esters (Formulae XXVI and XXVII) does not result in a decrease in FFA concentrations in rat serum compared to controls.

[0687] The data presented clearly show that DiLa esters lead to the release of large amounts of lactic acid (see Example 8), and therefore this compound should be useful in any (medical) situation where the ability to increase lactic acid is beneficial.

[0688] Furthermore, the FFA data for the DiLa ester compared to the data for LaKe and KeLa indicated that the decrease in FFA was primarily driven by the BHB component of LaKe versus KeLa.

[0689] DiLa esters may be important in the following scenarios:

[0690] -When one wishes to deliver lactic acid specifically.

[0691] - This compound can be combined (in different ratios) with other ketone esters such as 1,3-butanediol diacetoacetate, 1,3-butanediol dihexanoate, 1,3-butanediol, medium chain triglycerides, 3-hydroxybutyrate and (3R)-hydroxybutyl (3R)-hydroxybutyrate and / KeLA and LaKe to achieve dual delivery of BHB and lactate.

[0692] Example 16 - Chemoenzymatic Synthesis of L-(-)-Lactate Esters 3 (Formula XXII) and 4 (Formula XXIII) (LaKe Esters):

[0693] Purpose

[0694] Provided is the chemoenzymatic synthesis of L-(-)-lactates 3 and 4 (LaKe esters).

[0695] Materials and methods

[0696]

[0697] The procedure for the enzymatic synthesis of L-(-)-lactate 3 and 4 is as follows:

[0698] (R)(-)-1,3-Butanediol (1.50 g, 16.6 mmol, 1.0 eq.), (S)-(-)-ethyl lactate (19.7 g, 166 mmol, 10.0 eq.) and Cal B (1.50 g, 100 wt.%) (lipase B Candida antarctica immobilized on Immobead 150, recombinant from yeast (≈4000 U / g), SIGMA) were placed in a 50 mL flask and heated to 40°C. The mixture was kept at this temperature and stirred carefully (300-350 rpm) for 24 h. The mixture was cooled to room temperature and filtered under vacuum to remove the enzyme beads, and the filtrate was concentrated in vacuo. The crude product was subjected to FCC (pentane / EtOAc 1:1 to 1:2) to give the desired L-(-)-lactate (2.02 g, 75%; r. 3:4:20:1) as a clear oil.

[0699] result

[0700] NMR spectroscopy was used to confirm that the LaKe ester was produced as described above (data not shown).

[0701] in conclusion

[0702] L-(-)-Lactate esters 3 and 4 (LaKe esters) can be produced by chemoenzymatic synthesis. This provides an alternative method for preparing this compound that is shorter and more resource-friendly.

[0703] Example 17 - Chemoenzymatic Synthesis of L-(-)-Lactate Esters 7 (Formula XXVI) and 8 (Formula XXVII) (DiLa Esters):

[0704] Purpose

[0705] A chemoenzymatic synthesis of L-(-)-lactates 7 and 8 (DiLa esters) is provided.

[0706] Materials and methods

[0707]

[0708] The procedure for the enzymatic synthesis of L-(-)-lactates 7 and 8 is as follows:

[0709] (S)-(+)-1,2-propylene glycol (16.0 g, 0.21 mmol, 1.0 eq.), (S)-(-)-ethyl lactate (124 g, 1.05 mmol, 5.0 eq.) and Cal B 2 (16.0 g, 100 wt.%) (lipase B Candida antarctica immobilized on Immobead 150, recombined from yeast (≈4000 U / g), SIGMA) was placed in a 250 mL flask and heated to 40° C. The mixture was kept at this temperature and carefully stirred (300-350 rpm) for 24 h. The mixture was cooled to room temperature and filtered under vacuum to remove the enzyme beads, and the filtrate was concentrated in vacuo. The crude product was subjected to FCC (pentane / EtOAc 1:1 to 1:2) to obtain the desired L-(-)-lactate (16.1 g, 51%; r.7:8:10:1) as a clear oil.

[0710] result

[0711] NMR spectroscopy was used to confirm that the DiLa ester was produced as described above (data not shown).

[0712] in conclusion

[0713] L-(-)-Lactate esters 7 and 8 (DiLa esters) can be synthesized by a chemoenzymatic method. This provides an alternative method for preparing the compounds that is shorter and more resource friendly.

[0714] Without being bound by theory, the data presented in Examples 11 and 12 can also be applied to the synthesis of KeLa esters starting from (R)-(-)-ethyl 3-hydroxybutyrate and (S)-(+)-1,2-propanediol.

[0715]

[0716] Example 18 - Synthesis of R-(-)-β-Hydroxybutyrate Esters 14 and 15 (KeLa Esters) (Formulas XXIV and XXV)

[0717] Purpose

[0718] Synthesis of R-(-)-β-hydroxybutyrate esters 12 and 13 (KeLa esters):

[0719]

[0720] (R)-3-((tert-butyldimethylsilyl)oxy)butanoic acid ethyl ester (12):

[0721]

[0722] To a solution of (R)-(-)-3-hydroxybutyric acid ethyl ester (50.0 g, 378 mmol, 1.0 eq.) in anhydrous DCM (500 mL) was added TBSCl (59.8 g, 397 mmol, 1.05 eq.), followed by imidazole (51.5 g, 756 mmol, 2.0 eq.). The mixture was stirred at room temperature for 12 hours, then diluted with saturated NaCl solution and extracted with EtOAc. The organic layers were combined and washed with 5% HCl solution and saturated NaCl solution. The organic layer was then dried over Na2SO4, filtered and concentrated to give (R)-3-(tert-butyldimethylsilyloxy)butyric acid ethyl ester (89.0 g, 96%).

[0723] R f

[0724] 0.65 (pentane / Et2O 95:5, KMnO4).

[0725] 1 H NMR

[0726] (400MHz,CDCl3)δ H 4.31-4.21(m,1H),4.18-4.04(m,2H),3.93-3.84(m,1H),2.50-2.41(m,1H),2.39-2.30(m,1H),1.28 -1.22(t,J=7.3Hz,3H),1.21-1.16(d,J=6.22Hz,3H),0.87-0.83(s,9H),0.06-0.02(d,J=8.4Hz,6H).

[0727] 13 C NMR

[0728] (101MHz,CDCl3)δ C171.8,66.0,60.4,45.1,25.1,24.1,18.1,-4.4,-4.9.

[0729] IR (neat)

[0730] υ max / cm -1 2957,2930,2857,1738,1473,1376,1300,1255,1183,1139,1082.

[0731] HRMS

[0732] [M+Na] + =269.1543; found:269.1545.

[0733] [α] 24.0 D

[0734] -24.0°(C=10mg / mL,CHCl3).

[0735] (R)-3-((tert-Butyldimethylsilyl)oxy)butanoic acid:

[0736]

[0737] To a solution of (R)-3-((tert-butyldimethylsilyl)oxy)butanoate (89.0 g, 361 mmol, 1.0 eq.) in THF (500 mL) was added a cooled aqueous LiOH solution (469 mL, 1 M) at 0°C. The reaction mixture was stirred at 60°C overnight, then concentrated to half of the original volume and extracted with Et2O. The organic extracts were combined and then extracted with saturated aqueous NaHCO3. The aqueous layers were combined and acidified with 1M aqueous KHSO4 to a pH of ≈ 3-4. The aqueous solution was then thoroughly extracted with Et2O, and the organic layers were combined, dried over Na2SO4 and concentrated to give (R)-3-((tert-butyldimethylsilyl)oxy)butanoic acid (62.0 g, 79%) as an oil.

[0738] R f

[0739] 0.21 (pentane / EtOAc 4:1, KMnO4).

[0740] 1 H NMR

[0741] (400MHz,CDCl3)δ H4.32-4.23(m,1H),2.53-2.38(m,2H),1.24-1.18(d,J=6.2Hz3H),0.88-0.84(s,9H),0.08-0.04(d,J=6.9Hz,6H.

[0742] 13 C NMR

[0743] (101MHz,CDCl3)δ C 177.9,65.8,44.6,25.8,23.9,18.1,-4.4,-5.0.

[0744] IR(neat)

[0745] υ max / cm -1 3675,2958,2929,2900,2859,1710,1408,1379,1305,1253,1207,1133,1080.

[0746] HRMS

[0747] [M] - =217.1265; found:217.1262.

[0748] [α] 24.3 D

[0749] -16.5°(C=35mg / mL,CHCl3).

[0750] (S)-2-Hydroxypropyl (R)-3-hydroxybutyrate (14) and (S)-1-hydroxypropan-2-yl (R)-3-hydroxybutyrate (15):

[0751]

[0752] The acid (62 g, 284 mmol, 1.0 eq.), TBTU (101 g, 312 mmol, 1.1 eq.) and DIPEA (104 mL, 596 mmol, 2.1 eq.) were dissolved in anhydrous DMF (700 mL) and the mixture was stirred at room temperature for 1 hour. A solution of (S)-(+)-1,2-propylene glycol (23.5 mL, 312 mmol, 1.1 eq.) in anhydrous DMF (30 mL) was then added and the reaction mixture was stirred overnight at ambient temperature. The reaction mixture was diluted with CH2Cl2 and the resulting mixture was washed with 1M HCl aqueous solution, NaHCO3 aqueous solution and water in sequence. The organic layers were then combined, dried over Na2SO4, filtered and concentrated to give the crude product.

[0753] The crude TBS-ether was dissolved in THF:H2O:MeOH (2.5:2.5:1, 500 mL), KHSO4 (7.14 g, 52.5 mmol, 0.18 eq.) was added, and the mixture was stirred at room temperature overnight. MeOH was removed under reduced pressure, water was added, and the aqueous layer was then washed with Et2O. The aqueous phase was then thoroughly extracted with EtOAc (20x). The organic layer was collected, dried over Na2SO4, filtered and concentrated. The crude product was purified by FCC (pentane / EtOAc=1:1 to 1:2) to give the desired product (19.7 g, 44%; r.14:15:20:1).

[0754] R f

[0755] 0.28 (pentane / EtOAc 1:4, CAM).

[0756] 1 H NMR

[0757] 14:(400MHz,CDCl3)δ H 4.25-4.14(m,1H),4.13-4.07(dd,J=10.7Hz,2.6Hz,1H),4.05-3.97(m,1H),3.97-3.90(dd,J=10.7Hz,7.5Hz,1H), 3.57-4.34(s,1H),3.23-3.11(s,1H),2.53-2.38(m,2H),1.24-1.20(d,J=6.3Hz,3H),1.19-1.15(d,J=6.3Hz,3H).

[0758] 13 C NMR

[0759] 14:(101MHz,CDCl3)δ C172.7,69.7,65.8,64.6,43.3,22.8,19.1.

[0760] IR(neat)

[0761] υ max / cm -1 3368,2974,2934,1716,1377,1291,1178,1123,1072.

[0762] HRMS

[0763] [M+Na] + =185.0784; found:185.0788.

[0764] [α] 24.3 D

[0765] -17.4°(C=35mg / mL,CHCl3).

[0766] Example 19 - Chemical Synthesis of diLaKe

[0767] Purpose

[0768] Provide chemical synthesis of diLaKe

[0769] Materials and methods

[0770]

[0771] (S)-2-(Benzyloxy)propionic acid methyl ester (B)

[0772] Benzyl bromide (281 mmol, 33.6 mL) and silver (I) oxide (281 mmol, 65.2 g) were added to a solution of (S)-methyl lactate (225 mmol, 21.5 mL) in anhydrous dichloromethane (425 mL). The reaction mixture was stirred at room temperature under argon atmosphere for 72 hours. The reaction mixture was then The filtrate was filtered through a Celite pad and the filtrate was evaporated under reduced pressure (200 mbar) to give crude product A (43.15 g) as a colorless oil, which was used in the next step without further purification.

[0773] 1M KOH aqueous solution (225mmol, 225mL) was slowly added to a cooled solution of crude product A (43.15g) in ethanol (225mL), and the reaction mixture was stirred at 0°C for 30 minutes and then at room temperature for 1 hour. Then, the reaction mixture was concentrated to half volume under reduced pressure, and the resulting aqueous phase was washed with ether (2x 200mL). The aqueous phase was acidified with concentrated HCl aqueous solution (pHca.2) and then extracted with dichloromethane (2x200mL). The organic layers were combined, dried over anhydrous Na2SO4, filtered and evaporated under reduced pressure to give a colorless oil B (19.65g, 48%, 2 steps), which eventually crystallized as a white solid. Obtained 1 The H-NMR data were consistent with those reported in the reference (J. Nat. Prod. 2015, 78, 476-485).

[0774] Rf 0.48, tailing (Et2O / hexane 7:3, UV and KMnO4). 1 H-NMR (400MHz, CDCl3, δ): 7.42-7.28 (m, 5H), 4.70 (d, 1H; J = 11.4), 4.55 (d, 1H, J = 11.4), 4.12 (q, 1H, J = 6.8), 1.50 (d, 3H, J = 6.8).

[0775] (R)-Butane-1,3-diyl (2S, 2'S)-bis(2-(benzyloxy)propionate) (C)

[0776] To a solution of (S)-2-benzyloxypropionic acid B (109 mmol, 19.62 g) in anhydrous dichloromethane (340 mL) were added EDC hydrochloride (145 mmol, 27.84 g), triethylamine (145 mmol, 20.2 mL), DMAP (36.3 mmol, 4.44 g) and a solution of (3R)-butane-1,3-diol (36.3 mmol, 3.27 g) in anhydrous dichloromethane (20 mL) in sequence. The reaction mixture was stirred at room temperature for 20 hours under an argon atmosphere. The reaction mixture was then washed with a 1 M aqueous HCl solution (400 mL) and then with a saturated aqueous NaHCO3 solution (40 mL). The organic phase was dried over anhydrous Na2SO4, filtered and evaporated under reduced pressure. The resulting residue was purified by column chromatography (eluent 1: hexane / AcOEt 9:1, eluent 2: hexane / AcOEt 7:3) to give a colorless oil C (14.90 g, 99% yield).

[0777] Rf 0.68 (Et2O / hexane 1:1, UV and KMnO4). 1H-NMR (400MHz, CDCl3, δ): 7.44-7.27(m,10H),5.18-5.06(m,1H),4.69(d,1H,J=11.6),4.69(d,1H,J=

[0778] 11.6),4.45(d,1H,J=11.6),4.44(d,1H,J=11.6),4.30-4.21(m,1H),4.21-4.13(m,1H),4.07(q,1H,J=6 .8),4.04(q,1H,J=6.8),2.05-1.86(m,2H),1.45(d,3H,J=6.8),1.43(d,3H,J=6.8),1.30(d,3H,J=6.4). 13 C-NMR (100MHz, CDCl3, δ):173.3,172.9,137.7,128.6,128.6,128.1,128.1,128 .0,128.0,74.1,74.0,72.2,72.1,68.2,61.0,35.0,20.2,18.9,18.9.IR(neat,ν max / cm -1 ):1743,1140,1116.[α] D 26 -101.8(c=1.0,CHCl3).HRMS(ESI + ):m / z[M+Na] + calculated for C 24 H 30 NaO6437.1935, found 437.1948.

[0779] (R)-Butane-1,3-diyl (2S, 2'S)-bis(2-hydroxypropionate) (diLaKe)

[0780] Palladium on carbon (1.48 g, 10% Pd / C) was added to a solution of compound C (35.9 mmol, 14.88 g) in EtOH / AcOEt 1:1 (500 mL), and the reaction mixture was stirred at room temperature for 5 hours under a hydrogen atmosphere. The residue was filtered through a pad of Celite® (washed with dichloromethane) and the filtrate was evaporated under reduced pressure. The resulting residue was filtered through a pad of silica gel (eluent 1: Et2O / hexane 1:1, eluent 2: Et2O) to give diLaKe (7.90 g, 94% yield) as a colorless oil.

[0781] Rf 0.13 (Et2O / hexane 4:1, KMnO4). 1H-NMR (400MHz, CDCl3, δ):5.17-5.04(m,1H),4.33-4.15(m,4H),2.91-2.80(m,2H ),2.06-1.88(m,2H),1.41(d,3H,J=6.8),1.40(d,3H,J=7.2),1.30(d,3H,J=6.0). 13 C-NMR(100MHz,CDCl3,δ):175.7,175.4,69.3,66.9,66.8,61.7,34.8,20.5,20.4,20.1.IR(neat,ν max / cm -1 ):3442(br.),1729,1123.[α] D 23 -29.8 (c=1.0, CHCl3).HRMS(ESI + ):m / z[M+

[0782] Na] + calculated for C 10 H 18 NaO6257.0996, found 257.1004.

[0783] result

[0784] Using NMR spectroscopy and HRMS, it was confirmed that diLaKe was produced as described above.

[0785] in conclusion

[0786] diLaKe can be produced by chemical synthesis.

[0787] Example 20 - Chemoenzymatic Synthesis of diLaKe (Formula II)

[0788] Purpose

[0789] An alternative chemoenzymatic synthesis of diLaKe is provided.

[0790] Materials and methods

[0791]

[0792] (R)-Butane-1,3-diylbis(2-oxopropionate)(D)

[0793] To a mixture of pyruvic acid (400mmol, 35.2g) and (R)-1,3-butanediol (100mmol, 9.0g) in anhydrous toluene (700mL) was added p-toluenesulfonic acid monohydrate (6mmol, 1.1g). The reaction mixture was stirred at reflux for 22 hours and water was removed azeotropically (Dean-Stark). The reaction mixture was then cooled to room temperature. The solvent was evaporated under reduced pressure and the resulting residue was dissolved in dichloromethane (500mL) and washed with saturated NaHCO3 aqueous solution (2x 500mL). The organic layer was dried over anhydrous Na2SO4, filtered and evaporated under reduced pressure. The resulting residue was purified by column chromatography (AFCC; eluent 1: heptane / AcOEt 4:1, eluent 2: heptane / AcOEt 4:1 to 3:2) to give a pale yellow oil D (4.22g, 18% yield).

[0794] Rf0.48 (Et2O / hexane 4:1, KMnO4). 1 H-NMR (400MHz, CDCl3, δ): 5.21-5.10 (m, 1H), 4.37 (ddd, 1H, J = 11.2, J = 7.6, J = 5.2), 4.28 (ddd ,1H,J=11.2,J=6.0,J=6.0),2.47(s,3H),2.47(s,3H),2.20-2.00(m,2H),1.38(d,3H,J=6.4). 13 C-NMR(100MHz,CDCl3,δ):191.9,191.5,160.8,160.5,70.7,62.6,34.5,26.8,26.7,20.0.IR(neat,ν max / cm -1 ):1729,1143.[α] D 22 -24.6 (c=1.0, CHCl3).HRMS(ESI + ):m / z[M+Na] + calculated for C 10 H 14 NaO6253.0683, found 253.0683.

[0795] (R)-Butane-1,3-diyl (2S, 2'S)-bis(2-hydroxypropionate) (diLaKe)

[0796] A suspension of baker's yeast (YSC2, Sigma-Aldrich, 4.60 g) in distilled water (46 mL) was stirred at 25°C for 0.5 hours. A solution of compound D (1.00 mmol, 230 mg) in ethanol (4.6 mL) was then added, and the resulting suspension was stirred at 25°C for 2 hours. Dichloromethane (50 mL) was then added to the reaction mixture, and the entire mixture was vigorously stirred for 30 minutes. Some After that, the reaction mixture was vacuum filtered through filter paper and the phases were separated. The aqueous phase was extracted with dichloromethane (50 mL), and the combined organic layers were dried over anhydrous Na2SO4, filtered and evaporated under reduced pressure. The resulting residue was purified by column chromatography (AFCC; eluent 1: heptane / AcOEt 3:2, eluent 2: heptane / AcOEMt 3:2 to 3:7) to give diLaKe (68 mg, 29% yield) as a colorless oil.

[0797] What you get 1 H-NMR and 13 The C-NMR data as well as the HPLC trace matched those of diLaKe obtained by chemical synthesis.

[0798] result

[0799] NMR spectroscopy and HRMS were used to confirm that diLaKe was obtained by production as described above.

[0800] in conclusion

[0801] diLaKe can be produced by an alternative chemoenzymatic synthesis.

[0802] Example 21 - Animal Experiment and Quantification of BHB and Lactic Acid in Rat Serum - diLaKe (Formula II)

[0803] Materials and methods

[0804] As described in Examples 3 and 4, except that the values ​​were compared to a baseline (internal control). In addition, sampling was performed until 180 minutes.

[0805] result

[0806] like Figure 7 As shown, diLaKe caused an increase in serum levels of BHB and lactate. As expected, the increased serum concentrations of BHB and lactate both decreased again over time.

[0807] in conclusion

[0808] Here we show that treatment with diLaKe increases serum concentrations of both BHB and lactate in rats following oral administration.

[0809] Example 22 - Animal Experiments and Effects of DiLake Ester (Formula II) on Free Fatty Acids

[0810] Materials and methods

[0811] As described in Examples 3 and 5, except that the values ​​were compared to a baseline (internal control). In addition, sampling was performed until 180 minutes.

[0812] result

[0813] like Figure 8 As shown, diLaKe caused a decrease in FFA levels. As expected, the decreased FFA levels increased again over time.

[0814] in conclusion

[0815] Here, we show that oral administration of diLake resulted in a decrease in FFA concentrations in rat serum compared with a control group.

[0816] Example 23 - Effect of diLaKe ester (Formula II) on N-lactylphenylalanine (Lac-Phe)

[0817] Purpose:

[0818] It was demonstrated that the concentration of N-lactylphenylalanine (Lac-Phe) increased after oral administration of diLaKe.

[0819] Materials and methods:

[0820] As described in Example 21, except that N-lactylphenylalanine (Lac-Phe) was quantified using LC-MS / MS analysis.

[0821] LC-MS / MS analysis was performed as follows: 40 μL of rat plasma was diluted with 100 μL of water in a 2 mL tube, 115 μL of methanol (Merck hypergrade) and 420 μL of acetonitrile (Merck hypergrade) were added, vortexed, incubated at room temperature for 5 minutes, and then centrifuged at 10000×g for 5 minutes. 500 μL of supernatant was transferred to a 96-well plate (Eppendorf 1 mL deep well), evaporated to dryness at 30° C. under a stream of nitrogen, and then reconstituted in 100 μL of water containing 0.1% formic acid.

[0822] Pure calibration samples were prepared from the authentic standard compound (N,L-lactylphenylalanine) using the plasma sample preparation method at concentrations equivalent to 0, 1, 5, 10, 50, 100, 500 and 1000 μM in the original samples.

[0823] 7 μL of sample was injected into an ultra-high performance liquid chromatography system (Waters Acquity UPLC) coupled to a triple quadrupole mass spectrometer (Waters Xevo TQS) with a T3-UPLC column (Waters Acquity UPLC BEH T3, 1.8 μm, 100x2.1 mm) maintained at 40°C. The flow rate was 0.35 mL / min, with 100% mobile phase A (water with 0.1% formic acid) for 2 minutes, followed by a linear gradient transition to 40% B (methanol:acetonitrile 1:1, with 0.1% formic acid) (2-6 minutes), to 60% B (6-7 minutes), to 88% B (7-8 minutes) and to 100% B (9-10 minutes). At 11 minutes, the linear gradient returned to 100% A for 3 minutes for column equilibration, so the total run time was 14 minutes.

[0824] The samples were introduced into the mass spectrometer via electrospray negative ion mode with a capillary voltage of 2.2 kV, source temperature and desolvation temperature of 150 and 500 °C, respectively, and nitrogen flow rates of 50 L / h (cone) and 800 L / h (desolvation), respectively. The analytes were detected in multiple reaction monitoring mode with the following transitions (m / z) and cone voltage / collision energy (CV / CE): L-Lac-Phe (quantitative ion m / z: 236.2>88.1, CV / CE: 25 V / 15 eV; qualifier ion m / z: 236.2>147.1, CV / CE: 25 V / 15 eV). MassLynx 4.2 (Waters) was used for data acquisition and processing, including peak integration.

[0825] The calibration curve was constructed by linear regression of the peak area of ​​pure calibration samples with the nominal analyte concentration, based on eight points. The Lac-Phe concentration in rat plasma samples was derived based on its peak area and referenced to the calibration curve. The values ​​at different time points were compared with the baseline (internal control).

[0826] result:

[0827] like Fig. 9 As shown, we have shown that treatment with diLaKe (Formula II) results in an increase in serum concentrations of Lac-Phe in rats after oral administration. As expected, the increased Lac-Phe levels decrease again over time as the elevated levels of lactate also decrease.

[0828] in conclusion:

[0829] Oral administration of diLaKe (Formula II) results in increased serum concentrations of Lac-Phe.

[0830] Example 24 Synthesis of (R)-butane-1,3-diylbis(2-oxopropionate) (diPyKe) (Formula LXII)

[0831] Purpose

[0832] Provides the chemical synthesis of diPyKe.

[0833] Materials and methods

[0834] The synthesis of diPyKe is described in Example 20 above, ie, Compound D.

[0835] result

[0836] As shown in Example 20, the chemoenzymatic synthesis of diLake was first driven by the synthesis of the intermediate (R)-butane-1,3-diyl bis(2-oxopropionate) (Compound D), also referred to herein as Formula LXII or diPyKe.

[0837] Surprisingly, the reaction can be stopped at this intermediate, which itself is stable.

[0838]

[0839] in conclusion

[0840] We were able to obtain diPyKe, a novel compound suitable for oral administration.

[0841] Example 25 - Quantification of BHB and lactate in rat serum - diPyKe (Formula LXII)

[0842] Materials and methods

[0843] As described in Examples 3 and 4, except that the values ​​were compared to a baseline (internal control). In addition, sampling was performed up to 180 minutes and the experiment was performed with one rat.

[0844] result

[0845] like Fig.10 As shown, diPyKe caused an increase in serum levels of BHB and lactate. As expected, the increased serum concentrations of BHB and lactate decreased again over time.

[0846] in conclusion

[0847] Here we show that treatment with diPyKe results in increased serum concentrations of both BHB and lactate in rats following oral administration.

[0848] Example 26 - Effect of diPyKe ester (Formula LXII) on N-lactoylphenylalanine (Lac-Phe)

[0849] Purpose:

[0850] It was demonstrated that the concentration of N-lactylphenylalanine (Lac-Phe) increased after oral administration of diPyKe.

[0851] Materials and methods:

[0852] As described in Example 23, except that the experiment was conducted using one rat.

[0853] result:

[0854] like Fig.11 As shown, we have shown that treatment with diPyKe (Formula II) leads to increased serum concentrations of Lac-Phe in rats after oral administration. As expected, the increased Lac-Phe levels decrease again over time as the elevated levels of lactate also decrease.

[0855] in conclusion:

[0856] Oral administration of diPyKe (Formula LXII) resulted in increased serum concentrations of Lac-Phe.

[0857] References

[0858] 1-Sun,S.Li,H.,Chen,J.,Qian,Q.Lactic Acid:No Longer an Inert and End-Product ofGlycolysis.Physiology,2017,32,453-463.doi:

[0859] 10.1152 / physiol.00016.2017.

[0860] 2-Puchalska,P.,Crawford,P.A.Multi-dimensional Roles ofKetone Bodiesin Fuel Metabolism,Signaling,and Therapeutics.Cell Metab.201725,262-284.doi:.1016 / j.cmet.2016.12.022.

[0861] 3-Nielsen,R., N.,Gormsen,L,C.,Tolbod,L.P.,Hansson,N.H.,Sorensen,J.,Harms,H.J., J.,Eiskjaer,H.,Jespersen,N.R.,Mellemkjaer,S.,Lassen,T.R.,Pryds,K., H.E.,Wiggers,H.Cardiovascular Effects ofTreatmentWith the Ketone Body 3-Hydroxybutyrate in Chronic Heart FailurePatients.Circulation.2019,139,2129-2141.doi:10.1161 / CIRCULATIONAHA.118.036459.4-Thomsen,H.H.,Rittig,N.,Johannsen,M., A.B., J.O.,Jessen,N., N.Effects of3-hydroxybutyrate and free fattyacids on muscle protein kinetics and signaling during LPS-inducedinflammation in humans:anticatabolic impact ofketonebodies.Am.J.Clin.Nutr.2018,108,857-867.doi:

[0862] 10.1093 / ajcn / nqy170

[0863] 5-Lauritsen,K.M., E.,Svart,M., N.,Gormsen,L.C.Ketone Body Infusion Increases Circulating Erythropoietin and Bone MarrowGlucose Uptake.Diabetes Care.2018,41,e152-e154.doi:10.2337 / dc18-1421.

[0864] 6-M.C.Petersen,G.I.Shulman,Mechanisms ofInsulin Action and InsulinResistance.PhysiolRev.98,2133-2223(2018).

[0865] 7-Lambert K. Nikolaj F.Rittig,Emil F.Holmquist,Karl Anker

[0866] Jens Otto Lunde Niels Mogens Johannsen,Simultaneous determination ofβ-hydroxybutyrate andβ-hydroxy-β-methylbutyratein human whole blood using hydrophilic interaction liquid chromatographyelectrospray tandem mass spectrometry,Clinical Biochemistry,Volume 46,Issue18,2013,Pages 1877-1883.

[0867] 8-Rabinowitz&Sven Lactate:the ugly duckling ofenergymetabolism.Nature Metabolism volume 2,pages566–571(2020).

[0868] 9–Brooks,G.A.,Arevalo,J.A.,Osmond,A.D.,Leija,R.G.,Curl,C.C.andTovar.,A.P.Lactate in contemporary biology:A phoenix risen.J.Physiol.1-23(2021).10-Mayer,S.C.;Ramanjulu,J;Vera,M.D.;Pfizenmayer,A.J;Joullié,M.M,J.Org.Chem.,1994,59,5192-5205

Claims

1. Compounds of Formula II or Formula LXII or a pharmaceutically acceptable salt thereof.

2. A composition comprising one or more compounds according to claim 1.

3. The composition according to claim 2, comprising a compound of formula II.

4. The composition of claim 2, comprising a compound of formula LXII.

5. The composition according to any one of claims 2 to 4, further comprising one or more compounds selected from the following groups: Compound of formula X Compound of formula XI and Compound of formula XII or any pharmaceutically acceptable salt thereof, wherein R1, R2 and R3 are CH3, OH and H respectively, or R1, R2 and R3 are OH, H and CH3 respectively.

6. The composition according to any one of claims 2 to 5, further comprising one or more compounds selected from the following groups: Compound of formula XXII Compound of formula XXIII Compound of formula XXIV Compound of formula XXV Compound of formula XXVI Compound of formula XXVII Compound of formula XXVIII and Compound of formula XXIX or any pharmaceutically acceptable salt thereof.

7. A composition according to any one of claims 2 to 6, comprising a compound of formula II, a compound of formula XXII and / or a compound of formula XXIII.

8. The composition according to any one of claims 2-6, comprising a compound of formula LXII, a compound of formula XXII and / or a compound of formula XXIII.

9. A composition according to any one of claims 2 to 6, comprising a compound of formula II and at least one compound selected from 1,3-butanediol diacetoacetate, 1,3-butanediol dihexanoate, 1,3-butanediol, medium chain triglycerides, medium chain fatty acids, 3-hydroxybutyl 3-hydroxybutyrate and (3R)-hydroxybutyl (3R)-hydroxybutyrate.

10. A composition according to any one of claims 2 to 6, comprising a compound of formula LXII and at least one compound selected from 1,3-butanediol diacetoacetate, 1,3-butanediol dihexanoate, 1,3-butanediol, medium chain triglycerides, medium chain fatty acids, 3-hydroxybutyl 3-hydroxybutyrate and (3R)-hydroxybutyl (3R)-hydroxybutyrate.

11. The composition of any one of claims 2-10, wherein the composition comprises a compound of formula II and a compound of formula LXII.

12. A pharmaceutical composition comprising the compound of claim 1 or the composition of any one of claims 2-11.

13. A compound according to claim 1, a composition according to any one of claims 2 to 11 or a pharmaceutical composition according to claim 12 for use as a medicament.

14. The compound according to claim 1, the composition according to any one of claims 2 to 11, or the pharmaceutical composition according to claim 12, for use - Treat, prevent or alleviate inflammatory diseases, cancer, epileptic seizures, acute and chronic heart failure, cardiogenic shock, resuscitation, acidosis, traumatic brain injury, acute pancreatitis, hepatitis, myocardial infarction, atherosclerosis, ischemic disease, myocardial ischemia, pulmonary hypertension, systemic hypertension, organ transplant preparation, reperfusion injury after organ transplantation, burns, sepsis, dengue fever, cognitive dysfunction, neurodegeneration, oxidative stress, Alzheimer's disease, Parkinson's disease, multiple sclerosis, amyotrophic lateral sclerosis, epilepsy, astrocytoma, glioblastoma and Huntington's disease, sarcopenia, muscle atrophy, osteoporosis, wound healing dysfunction, damaged tissue, such as promoting tissue repair after injury, and infertility; - Treating, preventing or alleviating conditions caused by, exacerbated by or associated with elevated plasma free fatty acid levels in human or animal subjects; - Treating, preventing or alleviating a disease by increasing the plasma level of Lac-Phe in a human or animal subject; - Treat, prevent or alleviate viral infections, immunogenic conditions, non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), sarcopenia, loss of muscle / lean body mass, muscle fatigue, angina pectoris, extreme physical exertion, intermittent claudication, hypoxia, stroke, CNS hemorrhage and myocardial infarction, stress, obesity, diabetes, metabolic syndrome, autoimmune diseases, muscle damage or for improving motor function; and / or -Treat, prevent and / or reduce insulin resistance.

15. Use of the compound, composition or pharmaceutical composition according to claim 13 or 14, wherein the administration method is selected from oral administration, nasal administration, intradermal administration, intravenous administration, intramuscular administration, subcutaneous administration, intratracheal administration and transdermal administration.

16. Use of a compound, composition or pharmaceutical composition according to any one of claims 13 to 15, which is administered at a dose of 0.05-1.5 g / kg, preferably 0.15-1.5 g / kg, more preferably 0.2-1.0 g / kg, even more preferably 0.2-0.5 g / kg; Alternatively, it is administered at a dose of 0.05-15 g / kg, preferably 0.15-10 g / kg, more preferably 0.2-5 g / kg, even more preferably 0.2-2.5 g / kg, even more preferably 0.2-1.5 g / kg; Alternatively, it is administered at a daily dose of 0.05-45 g / kg, preferably 0.45-30 g / kg, more preferably 0.6-15 g / kg or 0.6-4.5 g / kg, even more preferably 0.6-1.5 g / kg.

17. Use of a compound, composition or pharmaceutical composition according to any one of claims 13 to 16, which is administered to a subject selected from the following groups: humans of all ages, mammals, including cattle, pigs, horses, sheep, goats, minks, ferrets, hamsters, cats, dogs; and / or birds, preferably the subject is a human.

18. A food ingredient comprising the compound of claim 1 or the composition of any one of claims 2-11.

19. A food product comprising the food ingredient of claim 18.

20. The food product according to claim 19, wherein the food product is selected from the group consisting of nutraceuticals, food supplements, dietary supplements, feeds, bars, candy bars, protein bars, powders, gels, beverages, drinks, yogurts, chewing gums, dairy products, sports drinks, confectionery products, ice cream, capsules, tablets, granules and sachets.

21. Use of a compound according to claim 1 or a composition according to any one of claims 2 to 11, - Reduction of circulating free fatty acids in the plasma of subjects in the non-therapeutic treatment of muscle damage, fatigue or improvement of motor function; - Suppress appetite, treat obesity, treat obesity non-therapeutically, promote weight loss, prevent, reduce and / or treat sarcopenia, maintain a healthy body weight or reduce the ratio of fat to lean muscle; - for non-therapeutic treatment of myocardial fatigue, skeletal muscle fatigue, and / or improvement of motor function, or for promoting alertness or improving cognitive function in a subject, or preventing neurodegeneration; - For promoting mental health; - For non-therapeutic increase in serum concentration of Lac-Phe; - Used to increase lifespan; - for promoting wound healing and tissue repair after injury, and / or -For promoting bone health, e.g. for the non-therapeutic treatment of osteoporosis.

22. A method for producing the compound of claim 1, the method comprising: a) providing 1,3-butanediol, such as (R)-1,3-butanediol and pyruvic acid; b) reacting the compound in step a in the presence of a catalytic acid, such as p-toluenesulfonic acid (pTsOH); c) obtaining a compound of formula LXII d) optionally, reacting the compound of formula LXII of step c) in the presence of a dehydrogenase, for example in the presence of baker's yeast; e) Optionally, if optional step d) is included, a compound of formula II is obtained 23. The method of claim 22, wherein steps d) and e) are not optional.

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