Quantification of renal acid-base excretion in people with short bowel

Abstract

The present invention relates to a method for determining a need for adjustment of parenteral supplementation, risk of developing acid-base imbalance, and monitoring development of acid-base imbalance in a subject receiving parenteral supplementation and having intestinal insufficiency, such as short bowel. In particular, the present invention relates to determining urinary net acid excretion (NAE) for evaluating acid-base imbalances in subjects having intestinal insufficiency, such as short bowel, in order to determine risk of developing and / or monitoring development of acid-base imbalance, as well as adjustment of parenteral supplementation.

Description

[0001] Quantification of renal acid-base excretion in people with short bowel

[0002] Technical field of the invention

[0003] The present invention relates to a method for determining a need for adjustment of parenteral supplementation, risk of developing acid-base imbalance, and monitoring development of acid-base imbalance in a subject receiving parenteral supplementation and having intestinal insufficiency, such as short bowel. In particular, the present invention relates to determining urinary net acid excretion (NAE) for evaluating acid-base imbalances in subjects having intestinal insufficiency, such as short bowel in order to determine risk of developing and / or monitoring development of acid-base imbalance as well as adjustment of parenteral supplementation.

[0004] Background of the invention

[0005] Intestinal failure is a life-threatening condition defined by reduced gut function below a minimum necessary for the absorption of macronutrients and / or water and electrolytes to maintain health and / or growth. As such, it requires parenteral support as organ replacement therapy. Short bowel as a consequence of surgical resection is the most common cause of intestinal failure and intestinal insufficiency. It is classified by the postoperative functional anatomy as end- jejunostomy (SB-J, formerly SB-type-1), jejunocolonic anastomosis with colon in continuity (SB-JC, formerly SB-type-2) and jejunoileocolonic anastomosis with colon in continuity (SB-JIC, formerly SB-type-3).

[0006] Metabolic acidosis due to intestinal bicarbonate loss is a well-known and common complication of high-volume diarrhea and of intestinal failure and intestinal insufficiency. Because of the difficulties measuring base losses in the stool the amount of intestinal bicarbonate (or other base) loss and thus the acidotic load that may manifest as metabolic acidosis has neither been quantified in detail nor characterized regarding its pathophysiologic origin.

[0007] Transport mechanisms for the absorption of nutrients as well as the absorption and secretion of electrolytes are segmentally distributed along the longitudinal axis of the small and large intestine. Thus, extensive resections of specific segments result in different types of malabsorption. Large amounts of water, sodium and potassium are often required in the parenteral admixture for intestinal failure patients to compensate intestinal losses and this can be guided by urinary sodium, serum potassium and the clinical assessment of volume status. In the management of intestinal failure and for the composition of parenteral support, venous blood gas analysis is normally used to monitor metabolic acid-base disturbances. However, this does not adequately reflect the acid load because of the unknown renal compensation of intestinal base losses.

[0008] Kato K, et al. The Latent Risk of Acidosis in Commercially Available Total Parenteral Nutrition (TPN) Products: a Randomized Clinical Trial in Postoperative Patients. J Clin Biochem Nutr. 2009 Jul;45(l):68-73') discloses calculation of blood acid-base status by using standard automated laboratory techniques. Urine is collected for the measurements of urine electrolytes, pH, creatinine, and urinary net acid excretion (NAE). A formula for calculating urinary NAE being NAE = NH4+excretion + TA excretion - HCOs" excretion is disclosed. Further, it is disclosed that the NAE can be used for evaluating the risk of metabolic acidosis or alkalosis in patients with resected bowel receiving commercial total parenteral nutrition (TPN). However, comparison of calculated NAE with reference values for determining the degree of acid-base disturbances or the degree of acidosis or alkalosis for individual adjustment of parenteral nutrition is not disclosed.

[0009] US 2013 / 129838 Al discloses methods for reducing acid-base imbalance in subjects undergoing parenteral supplementation by adjusting formulations. The method comprises adjusting amino acid levels in the parenteral supplementation and adjusting acid-base components. However, there is no disclosure of regularly determining and monitoring NAE in subjects having short bowel and receiving parenteral supplementation for a permanent and long-lasting period.

[0010] Hence, an improved method for determining the risk of developing and / or degree of acid-base imbalance in short bowel patients would be advantageous, and in particular a more efficient and / or reliable method for determining the risk of developing and / or degree of acid-base imbalance in short bowel patients in order to determine the need of adjustment of parenteral supplementation would be advantageous. Furthermore, an improved use of urinary net acid excretion (NAE) for evaluating acid-base balance disturbances or imbalances in subjects having intestinal insufficiency in order to determine the risk of developing and / or monitoring development of acidic imbalance or alkaline imbalance in said subjects would be advantageous, and in particular a more efficient and / or reliable use of urinary net acid excretion (NAE) for adjustment of parenteral supplementation in subjects having intestinal insufficiency and receiving parenteral supplementation would be advantageous.

[0011] In summary, it is suggested that the net acid / base load of the individual parenteral supplementation must become adjusted so that subjects / patients display full normalization of systemic acid-base parameters and reach urine NAE within the normal physiological range and does not display alkaline or acidic imbalance as defined in the invention.

[0012] Summary of the invention

[0013] The acid / base components (NH4+, HCOs", titratable acids (TA) and pH) have been measured in urine samples from patients with intestinal failure or short bowel and receiving parenteral support. Based on the measured biomarker and the 24-hour diuresis, excretion rates were calculated. Subsequently, net acid excretion was calculated (NAE = NH4+excretion + TA excretion - HCOs" excretion). When comparing NAE of short bowel patients with NAE from healthy controls striking differences were observed. Some patients display NAE above the 95% percentile of the controls. These patients therefore have a condition where a very large acid load provides a systemic burden that interferes with their health (acidic imbalance or even acidosis). Other patients are in the opposite situation, where they excrete more base in the urine (alkaline imbalance or even alkalosis) than 5% of the healthy controls. Alkaline imbalance or alkalosis likely adds extra strain on the organism, which reduces their health.

[0014] Example 1 shows a characterization of the short bowel patient cohort. Example 2 shows venous base excess, serum anion gap, and total provision of base equivalents from all patients in the cohort. Further, venous base excess and acid-base measurements from 24h urine collections for each type of SB-anatomy are shown.

[0015] Example 3 shows parenteral base equivalents and its impact on base excess. SB- JC patients had a small but significantly acidic base excess indicating underbuffering, while SB-J patients had a slightly positive base excess, indicating too intense buffering.

[0016] Example 4 shows the role of the residual small intestine for total acid load. Small bowel length is not significantly different between SB-J and SB-JC and small bowel length is not related to intestinal base loss in either SB-J nor in SB-JC or SB-JIC. Taken together, this indicates that this base loss is brought about by the presence of colon in continuity specifically mid-distal colon in continuity.

[0017] Example 5 shows the association between parenteral support components, such as acetate, chloride, and sodium, and acid-base-parameters.

[0018] Example 6 shows the identification of the predetermined reference range for urine NAE in healthy subjects thereby demonstrating the normal physiological range as defined in the invention.

[0019] Example 7 shows a characterization of the acid-base parameters of the short bowel patient cohort.

[0020] Thus, an object of the present invention relates to an improved method for determining a need for adjustment of parenteral supplementation, risk of developing acid-base imbalance, and monitoring development of acid-base imbalance in a subject receiving parenteral supplementation and having intestinal insufficiency, such as short bowel.

[0021] In particular, it is an object of the present invention to provide a method that solves the above mentioned problems of the prior art with an improved method for determining a need for adjustment of parenteral supplementation, for determining the risk of developing acid-base imbalance, and for monitoring the development of acid-base imbalance in a subject receiving parenteral supplementation and having intestinal insufficiency.

[0022] Thus, one aspect of the invention relates to a method for determining a need for adjustment of parenteral supplementation in a subject receiving parenteral supplementation and having short bowel, the method comprising providing a urine sample from said subject; measuring levels of at least three biomarkers in said urine sample;

[0023] - determining / calculating a net acid excretion (NAE) value based on the levels of the at least three biomarkers;

[0024] - comparing the NAE value with a reference range having an upper and a lower limit; wherein, if the NAE value exceeds the upper limit or if the NAE value is below the lower limit, it is indicative of said subject needing adjustment of parenteral supplementation, or wherein, if the NAE value is within the reference range, it is indicative of said subject not needing adjustment of parenteral supplementation.

[0025] A second aspect of the present invention relates to a method for monitoring risk of developing acid-base imbalance in a subject receiving parenteral supplementation and having short bowel, the method comprising providing a urine sample from said subject; measuring levels of at least three biomarkers in said urine sample;

[0026] - determining / calculating a net acid excretion (NAE) value based on the levels of the at least three biomarkers;

[0027] - comparing the NAE value with a reference range having an upper and a lower limit; wherein, if the NAE value exceeds the upper limit, it is indicative of said subject being at risk of developing acidic imbalance, or wherein, if the NAE value is below the lower limit, it is indicative of said subject being at risk of developing alkaline imbalance, or wherein, if the NAE value is within the reference range, it is indicative of said subject not being at risk of developing acid-base imbalance.

[0028] Yet a third aspect of the present invention is to provide a method for monitoring development of acid-base imbalance in a subject receiving parenteral supplementation and having short bowel, the method comprising providing a first urine sample from said subject before said subject receives adjustment of parenteral supplementation;

[0029] - adjusting the parenteral supplementation after said first urine sample has been provided; providing a second urine sample wherein the second urine sample has been obtained after the subject has received the adjusted parenteral supplementation; measuring levels of at least three biomarkers in the first and the second urine sample;

[0030] - determining / calculating a net acid excretion (NAE) based on the levels of the at least three biomarkers in the first and the second urine sample;

[0031] - comparing the NAE value with a reference range having an upper and a lower limit in the first and the second urine sample; wherein, the difference between the NAE value in the first or the second urine sample exceeding the upper limit and the upper limit is indicative of a degree of acidic imbalance in the subject, or wherein, the difference between the NAE value in the first or the second urine sample below the lower limit and the lower limit is indicative of a degree of alkaline imbalance in the subject, or wherein, if the NAE value in the first or the second urine sample is within the reference range, it is indicative of said subject not having acid-base imbalance.

[0032] Still fourth aspect of the present invention is to provide use of a net acid excretion (NAE) value for determining a need for adjustment of parenteral supplementation in a subject receiving parenteral supplementation and having short bowel. A fifth aspect of the present invention is to provide use of a net acid excretion (NAE) value for monitoring risk of developing acid-base imbalance in a subject receiving parenteral supplementation and having short bowel.

[0033] A sixth aspect of the present invention is to provide use of a net acid excretion (NAE) value for monitoring development of acid-base imbalance in a subject receiving parenteral supplementation and having short bowel.

[0034] Brief description of the figures

[0035] Figure 1 shows dot plots for base excess (A), serum anion gap (B), and parenteral provision of base equivalents (C). Venous base excess was 0.6 ± 3.9 mmol / l (mean ± standard deviation), serum anion gap was 14.1 ± 1.5 mmol / l, and parenteral provision of base equivalents was 66 ± 42 mmol / day,

[0036] Figure 2 shows venous base excess and acid-base measurements from 24h urine collections for each type of SB-anatomy. Observations: 48 (urine measurements) and 64 (base excess). Patients: 34 (urine measurements) and 44 (base excess),

[0037] Figure 3 shows (A) Daily base infusion for each anatomical SB-type. (B) Estimated total acid load (48 observations from 34 patients). Adding daily amount of infused base equivalents to NAE still results in significantly higher need for acid excretion in SB-JC compared to SB-J. (C) Base excess (BE) correlates with estimated total acid load. There is no interaction between anatomy type and BE and total acid load (or NAE). The association was fitted with a pooled regression line. Black circle: SB-J, light grey circle: SB-JC, dark circle: SB-JIC.

[0038] Figure 4 shows association of small bowel length with base excess (A+B), base infusion (C+D) and total acid load (E+F). Black circle: SB-J. Light grey circle: SB- JC, Black square: SB-JIC.

[0039] Figure 5 shows the association of NAE with parental supplementation. Net acid excretion (NAE) was negatively correlated to parenteral support of sodium (A), acetate (B), and chloride (C); but not with acetate / chloride ratio (D). Black circle: SB-J, light grey circle: SB-JC, dark grey circle: SB-JIC. Figure 6 shows the 24h NAE in 46 patients with intestinal failure and 25 healthy control individuals.

[0040] Figure 7 shows an illustration of the different short bowel (SB) classifications.

[0041] The present invention will now be described in more detail in the following.

[0042] Detailed description of the invention

[0043] Definitions

[0044] Prior to discussing the present invention in further details, the following terms and conventions will first be defined:

[0045] Parenteral supplementation

[0046] Parenteral supplementation is a way for a subject to receive nutrients by bypassing the digestive system of said subject. Parenteral supplementation can be supplementary (partial) or complete (total parenteral nutrition). Thus, parenteral supplementation as used herein means feeding intravenously (through a vein). "Parenteral" means "outside of the digestive tract." Accordingly, parenteral nutrition / supplementation bypasses the entire digestive system. Certain medical conditions may require parenteral nutrition / supplementation for a short or longer time or even for life.

[0047] Parenteral nutrition / supplementation as used herein refers to a chemical formula with standard variations and can be customized to specific nutritional requirements. It may include different amounts of any of the six essential nutrients that the body requires: water, carbohydrates, proteins, fats, vitamins and minerals.

[0048] "Parenteral supplementation" may be used interchangeably with "parenteral nutrition".

[0049] In an embodiment of this invention, the parenteral supplementation is partial. In a preferred embodiment of this invention, the parenteral supplementation is complete (total parenteral nutrition). Adjustment of parenteral supplementation

[0050] Parenteral supplementation as mentioned above can be customized / adjusted to specific nutritional requirements of patients in need thereof such as patients with intestinal insufficiency and / or intestinal failure. However, the pathophysiology and compliance of patients receiving parenteral supplementation as well as pharmacy processing may lead to acid-base imbalance in the patients. In order to counteract such imbalances, adjustment of parenteral supplementation as used herein is performed. Hence, there is a need to adjust one or more components of the parenteral supplementation in a feasible way to balance acid-base homeostasis in patients with e.g. intestinal insufficiency and / or intestinal failure.

[0051] The adjustment of parenteral supplementation may also comprise administration of one or more organic ions such as acetate, lactate, malate, and / or chloride, or one or more amino acids, such as cysteine, methionine, lysin, arginine, histidine, glutamic acid, and / or aspartic acid. Thus, adjustment of parenteral supplementation may also include acid / base adjustments. Cysteine, methionine, lysine, arginine and histidine all form acids when metabolized. Conversely, glutamic acid and aspartic acid form bases when metabolized. If a subject has too much acid load, the acid load can be reduced by adding less cysteine, methionine, lysine, arginine and histidine and / or by increasing the amount of glutamic acid and aspartic acid. If the patient is base loaded, the reverse will be required i.e. the base load can be reduced by adding more cysteine, methionine, lysine, arginine and histidine and / or by decreasing the amount of glutamic acid and aspartic acid.

[0052] An embodiment of the invention relates to a method for determining a need for adjustment of amino acid levels in parenteral supplementation in a subject receiving parenteral supplementation and having intestinal insufficiency, such as short bowel, the method comprising providing a urine sample from said subject; measuring levels of at least three biomarkers;

[0053] - determining / calculating a net acid excretion (NAE) value based on the levels of the at least three biomarkers;

[0054] - comparing the NAE value with a reference range having an upper and a lower limit; wherein, if the NAE value exceeds the upper limit or if the NAE value is below the lower limit, it is indicative of said subject needing adjustment of amino acid levels in said parenteral supplementation, or wherein, if the NAE value is within the reference range, it is indicative of said subject not needing adjustment of amino acid levels in said parenteral supplementation.

[0055] Net acid / base load

[0056] As used herein the "net acid-base load" refers to the acid-base load that is present in a given parenteral supplementation administered to a subject in need thereof. Dietary acid-base load is normally a balance between hydrogen iongenerating foods such as meats, cereals and dairy, and foods providing base precursors such as fruits and vegetables. Healthy renal function allows excretion of excess dietary hydrogen ions, which is thus reflected in urine pH.

[0057] An embodiment of the invention relates to a method for determining a need for adjustment of net acid / base load in parenteral supplementation in a subject receiving parenteral supplementation and having intestinal insufficiency, such as short bowel, the method comprising providing a urine sample from said subject; measuring levels of at least three biomarkers in said urine sample;

[0058] - determining / calculating a net acid excretion (NAE) value based on the levels of the at least three biomarkers;

[0059] - comparing the NAE value with a reference range having an upper and a lower limit; wherein, if the NAE value exceeds the upper limit or if the NAE value is below the lower limit, it is indicative of said subject needing adjustment of net acid / base load in said parenteral supplementation, or wherein, if the NAE value is within the reference range, it is indicative of said subject not needing adjustment of net acid / base load in said parenteral supplementation. Intestinal insufficiency

[0060] Intestinal insufficiency as used herein refers to the reduction of gut absorptive function that does not require intravenous supplementation to maintain health and / or growth. In other words, intestinal insufficiency is the compensated form of short bowel in which oral autonomy is maintained and which often requires enteral supplementation of electrolytes and macronutrients. Enteral nutrition, also known as tube feeding, is a way of sending nutrition right to the stomach or small intestine.

[0061] Transport mechanisms for the absorption of nutrients as well as the absorption and secretion of electrolytes are segmentally distributed along the longitudinal axis of the small and large intestine. Thus, extensive resections of specific segments result in different types of malabsorption. Short bowel as a consequence of surgical resection is the most common cause of intestinal insufficiency and intestinal failure.

[0062] Intestinal failure

[0063] Intestinal failure (IF) as used herein refers to the inability of the gut to absorb necessary water, macronutrients (carbohydrate, protein, and fat), micronutrients, and electrolytes sufficient to sustain life and requiring intravenous supplementation / parenteral supplementation) or replacement to maintain health or facilitate growth. Acute IF (types 1 and 2) is the initial phase of the illness and may last for weeks to a few months, and chronic IF (type 3) from months to years.

[0064] IF may be categorized into:

[0065] • Type 1, which is usually self-limiting and typical examples include postoperative ileus.

[0066] • Type 2, which presents following intra-abdominal surgery with metabolically unstable patients often with hostile abdomens, fistulae, or adhesions.

[0067] • Type 3, which is chronic IF and includes patients who have progressed from instability to stability, requiring long-term intravenous management of their IF over months to years.

[0068] IF can be further classified according to pathophysiology: short bowel syndrome (SBS), intestinal fistulae, dysmotility, mechanical obstruction, and extensive small bowel mucosal disease. Short bowel

[0069] Short bowel as a consequence of surgical resection is the most common cause of intestinal failure and intestinal insufficiency. It is classified by the postoperative functional anatomy as end-jejunostomy (SB-J, formerly SB-type-1), jejunocolonic anastomosis with colon in continuity (SB-JC, formerly SB-type-2) and jejunoileocolonic anastomosis with colon in continuity (SB-JIC, formerly SB-type- 3).

[0070] Short bowel is a permanent condition, hence, a subject or a patient having short bowel requires long-term parenteral supplementation i.e. lifelong parenteral supplementation. Therefore, said subject having short bowel and receiving longterm parenteral supplementation requires monitorization on a regular basis to determine risk of developing acid-base imbalance due to continuous administration of parenteral supplementation and thereby determining any need for adjustment of parenteral supplementation. In that way, the subject will constantly maintain systemic acid-base parameters within the normal physiological range.

[0071] Subject

[0072] "Subject" as described herein is to be understood as an animal or a human being, said human being including healthy individuals and individuals with acid-base imbalance (i.e. subjects receiving parenteral supplementation), where healthy individuals are to be understood as individuals not suffering from acid-base imbalance.

[0073] 24h urine sample or 8 hour urine sample

[0074] "24h urine sample" as used herein refers to a 24-hour urine sample that is done by collecting the urine of a subject in a special container over a full 24-hour period. An "8 hour urine sample as used herein refers to an 8-hour urine sample that is done by collecting the urine of a subject in a special container over a full 8- hour period.

[0075] Spot urine sample

[0076] "Spot urine sample" as used herein refers to urine samples collected at one-time point during the day. This also means "simple spot urine samples". In an embodiment of the invention, spot urine samples are collected over preferably 1-2 hours, such as more preferably over 1 hour, such as most preferably over 30 minutes. First and second urine sample

[0077] As used herein, a "first urine sample" is a urine sample from a subject prior to adjustment of parenteral supplementation. A "second urine sample" is a urine sample obtained at a later time point than the first urine sample and after adjustment of parenteral supplementation. The NAE values obtained from the first urine sample and second urine sample are to be compared with each other and with a reference range in order to determine a degree of acid-base imbalances / disturbances, such as acidosis or alkalosis.

[0078] A certain amount of time, e.g. 12-24 hours must pass from the time the parenteral supplementation has been administered / given to the subject and / or adjustment of the parenteral supplementation has been administered / given to the subject before a first and / or a second urine sample is provided, respectively.

[0079] "A certain amount of time" is to be understood as about 4-8 hours, such as about 4-12 hours, such as about 4-24 hours, such as about 4-36 hours, such as preferably about 8-12 hours, such as more preferably about 8-24 hours, such even more preferably about 8-36 hours, such likewise even more preferably about 12-36 hours, and such as the most preferable about 12-24 hours.

[0080] The amount of time that passes after administration of the parental supplementation and the first urine sample collection can be different from the amount of time that passes after administration of the adjusted parental supplementation and the second urine sample collection.

[0081] A first or a second urine sample is to be understood as a single urine sample obtained at a certain point in time, or two or more urine samples obtained over a given time period such as two or more samples obtained over at least three hours.

[0082] "Ammonium" and "bicarbonate"

[0083] As used herein, "ammonium" is used interchangeably with "NH4+" and "bicarbonate" is used interchangeably with "HCO3 ".

[0084] Titratable acid (TA)

[0085] The acid in human urine is bound in buffers such as phosphates and other inorganic anions derived from normal metabolism. "Titratable acid (TA)" or "urinary TA / UTA" as used herein thus refers to protons bound to these anions. Biomarker

[0086] Biomarkers are measurable indicators of some biological state or condition, hence they are objective, quantifiable characteristics of biological processes.

[0087] As used herein "biomarker" refers to a general term representing acid-base components, for example NH4+, HCOs", titratable acids (TA) or pH. In a preferred embodiment, the biomarkes are one or more of NH4+, HCOs", titratable acids (TA) and / or pH. In an even more preferred embodiment, the biomarkers are one or more of NH4+, HCOs" and titratable acids (TA).

[0088] Acid- base imbalance

[0089] Acid-base imbalance is an abnormality of the human body's normal balance of acids and bases that causes the plasma pH to deviate out of the normal range (7.35 to 7.45). The acid-base imbalance can be due to retention of acids or bases. Acid-base disturbance or imbalance provokes automatic compensatory mechanisms that push the blood pH back toward the normal range. Thus, urine samples are a better indicator of any acid-base imbalance, as the acid-base fluctuations in urine are not affected by the compensatory mechanisms as seen in blood samples.

[0090] Acidic imbalance and alkaline imbalance

[0091] As used herein "acidic imbalance" refers to acid-base imbalance where the pH of the urine is acidic, whereas "alkaline imbalance" refers to acid-base imbalance where the pH of the urine is alkaline.

[0092] Compound

[0093] "Compound" as described herein is to be understood as a drug, medicament or other chemical compound, that may be used for treating subjects having acid / base imbalance or that may be used to decrease or eliminate the risk of developing acid-base imbalance. This may be approved drugs already in use or new compounds, which are currently being tested or developed for the purpose of treating subjects having acid / base imbalance.

[0094] According to the invention one or more compounds can be administered to subjects to decrease or eliminate the risk of developing acid-base imbalance, such as acidosis or alkalosis. As used herein, said compounds may be selected from the group consisting of: proton pump inhibitors to reduce production of stomach acid by glands in the lining of the stomach,

[0095] - acid binders or proton chelators, such as potassium binders, to remove or reduce the amount of acid in the gastrointestinal tract, thereby preventing intestinal absorption of acids, and / or minerals, such as chloride, magnesium, potassium and / or sodium.

[0096] Acidosis

[0097] Acidosis is a condition in which there is an acidic imbalance in the body fluids i.e. too much acid in the body fluids. The kidneys and lungs maintain a proper pH level and balance of chemicals called acids and bases in the body. Acidosis occurs when acid builds up or when bicarbonate (a base) is lost. Acidosis is classified as either respiratory or metabolic acidosis.

[0098] Respiratory acidosis develops when there is too much carbon dioxide (an acid) in the body. This type of acidosis is usually caused when the body is unable to remove enough carbon dioxide through breathing.

[0099] Metabolic acidosis can develop if there is too much acid in the blood that wipe out bicarbonate (high anion gap metabolic acidosis) or if too much bicarbonate is lost in the blood as a result of kidney disease or kidney failure (normal anion gap metabolic acidosis). Metabolic acidosis also occurs when the body produces too much acid, or the kidneys do not remove enough acids from the blood.

[0100] The causes of metabolic acidosis may include diabetes-related acidosis, hyperchloremic acidosis, lactic acidosis, renal tubular acidosis, intestinal insufficiency and intestinal failure.

[0101] Further, acidosis may be classified as subclinical acidosis or overt acidosis. Subclinical acidosis is a milder degree of acidosis, whereas overt acidosis is a more severe form of acidosis. Acidosis is commonly determined in a blood sample.

[0102] Subclinical acidosis

[0103] The term "subclinical acidosis" is to be understood as a disorder that is below the limit of clinical detection. Subclinical acidosis is a milder degree of acidosis.

[0104] Alkalosis

[0105] Alkalosis is a condition in which there is an alkaline imbalance in the body fluids i.e. the body fluids have excess base (alkali). The kidneys and lungs maintain the proper pH level and balance of chemicals called acids and bases in the body. Decreased carbon dioxide (an acid) level or increased bicarbonate (a base) level makes the body too alkaline, a condition called alkalosis. Alkalosis is commonly determined in a blood sample.

[0106] Respiratory alkalosis is caused by a low carbon dioxide level in the blood. Metabolic alkalosis is caused by too much bicarbonate in the blood. It can also occur due to certain kidney diseases. Metabolic alkalosis may include hypochloremic alkalosis when there's a significant decline of chloride in the body and hypokalemic alkalosis when the body lacks the normal amount of potassium. Further, alkalosis may be classified as subclinical alkalosis or overt alkalosis.

[0107] Subclinical alkalosis is a milder degree of alkalosis, whereas overt alkalosis is a more severe form of alkalosis.

[0108] Subclinical alkalosis

[0109] The term "subclinical alkalosis" is to be understood as a disorder that is below the limit of clinical detection. Subclinical alkalosis is a milder degree of alkalosis.

[0110] Anion gap

[0111] An anion gap is the difference between the positive and negative electric charges in the electrolytes measured in the blood. Often, only chloride, potassium and sodium are measured in the blood. As there are often fewer Cl- than the sum of K+and Na+in the blood, a difference occurs and is called an anion gap. This gap is often filled by HCO3 ). Electrolytes are ions that help regulate many metabolic processes in the body, such as bringing nutrients into cells and taking waste products out of cells. Examples of electrolytes include sodium, calcium, potassium, chloride, bicarbonate, and phosphate.

[0112] Blood is electroneutral (equal numbers of cations and anions), but you often only measure chloride of the anions, whereas you measure potassium and sodium of the cations. There are often fewer Cl- than the sum of K+ and Na+, and the lack of negative ions so that cat- and anions are in equal weight is called an anion gap (and basically it is HCO3- that fills the anion gap). So the larger the tank's anion gap, the more HCO3-, i.e. the more base is present)

[0113] Net acid excretion

[0114] Net acid excretion (NAE) is the net amount of acid excreted in the urine per unit time. Hence, NAE provides a precise estimate of endogenous acid production in the body. The net acid excretion in the urine is equal to the sum of the titratable acids (TA) and NH4+minus HCOs". The various acid-base components can be used to determine or calculate the NAE, said acid-base components may comprise NH4+, HCOs", titratable acids (TA), citrate, pH or any combination thereof. However, preferably the acid-base components used for calculating or determining NAE are NH4+, HCOs" and titratable acids (TA).

[0115] Measurement of urinary net acid excretion (NAE) provides a sensitive and clinically useful method for evaluating acid-base balance.

[0116] Determining / calculating NAE

[0117] As used herein, the NAE value is obtained in vitro by measuring or sensing acid / base components in urine samples or the relationship between measured or sensed acid / base parameters in urine samples. By "measuring" or "sensing" is also meant "determining" values in the urine. In some embodiments, calculating values in the urine may also be an option. The measured or sensed parameters may be at least one or more of the biomarkers urinary NH4+, HCOs", titratable acids (TA), citrate and / or pH.

[0118] Reference range

[0119] A reference range is obtained by determining the urine NAE of healthy control individuals and thereby obtaining an interval having an upper and lower limit. The upper limit may be the higher 95% percentile of controls and the lower limit may be the lower 5% percentile of controls, and thereby obtaining a 95% prediction interval.

[0120] The reference range may be a predetermined reference range. Preferably, said predetermined reference range or reference range has an upper limit of 95% percentile of controls and a lower limit of 5% percentile of controls. Surprisingly, this predetermined reference range or reference range can be used to identify subjects having acid-base disturbances or imbalances.

[0121] Subject receiving parenteral supplementation

[0122] A subject receiving parenteral supplementation is a patient having a degree of intestinal insufficiency or even intestinal failure, which results in an inability or impaired ability to naturally absorb nutrients and electrolytes. Hence, said subject has reduced gut function below a minimum necessary for the absorption of macronutrients, water and / or electrolytes to maintain health and / or growth. Thus, the subject requires parenteral support as organ replacement therapy. Accordingly, as used herein "receiving parenteral supplementation" means that a subject has been administered or is administered parenteral supplementation. eGFR

[0123] "eGFR" as used herein refers to estimated glomerular filtration rate. eGFR is an estimated number based on the blood level of creatinine and the humans age, sex of the respective patient. eGFR is cost and time-effective assessment of GFR but lack accuracy as compared to mGFR.

[0124] "Software / "AI"

[0125] Another embodiment of the present invention relates to processor system programmed to operate according to a machine learning (ML) algorithm for determining a need for adjustment of parenteral supplementation, such as adjustment of net acid / base load or amino acid levels in said parenteral supplementation, monitoring risk of developing acid-base imbalance, and / or monitoring development of acid-base imbalance based on a relationship between the levels of biomarkers of the invention, the machine learning (ML) algorithm being trained, and / or being trainable, on data obtained by a method according to the first aspect of the invention or the second aspect of the invention or the third aspect of the invention. Preferably, said biomarkers of the invention are three biomarkers, even more preferably said three biomarkers are NH4+, HCOs", and titratable acids (TA).

[0126] In yet another embodiment, the invention relates to use of a machine learning (ML) algorithm trained on data obtained by a method according to the first or second or third aspect of the invention to determine a need for adjustment of parenteral supplementation, such as adjustment of net acid / base load or amino acid levels in said parenteral supplementation, monitor risk of developing acidbase imbalance, and / or monitor development of acid-base imbalance based on a relationship between the levels of biomarkers of the invention. Preferably, said biomarkers of the invention are three biomarkers, even more preferably said three biomarkers are NH4+, HCOs", and titratable acids (TA).

[0127] An embodiment of the present invention relates to a system suitable for executing an algorithm (such as machine learning (ML) algorithm) for determining a need for adjustment of parenteral supplementation, such as adjustment of net acid / base load or amino acid levels in said parenteral supplementation, monitoring risk of developing acid-base imbalance, and / or monitoring development of acidbase imbalance based on a relationship between the levels of biomarkers of the invention, the (machine learning) system being trained, and / or being trainable, on data provided according to the first, second or third aspect of the invention.

[0128] Advantageously, the invention may also relate to a method for training a machine learning (ML) system for determining a need for adjustment of parenteral supplementation, such as adjustment of net acid / base load or amino acid levels in said parenteral supplementation, monitoring risk of developing acid-base imbalance, and / or monitoring development of acid-base imbalance based on a relationship between the levels of biomarkers of the invention, such as according to the first, second or third aspect of the invention.

[0129] Thus, yet an embodiment of the invention relates to a method comprises the steps of:

[0130] -receiving training data comprising a first set of information (1SI), such as a first database, and a second set of information (2SI), such as a second database, -training the system for estimating score variants using said training data, and

[0131] - validating the system using correlated specific score variants to the frequency information, such as frequency of the variant.

[0132] Preferably the system and / or algorithm and / or method is implemented on a computer, thus being computer-implemented.

[0133] Below is given a list of some no-limiting types of algorithms that are particularly suited for machine learning (ML) system and / or training of a (ML) system using urine data:

[0134] 1. Deep Learning Algorithms: Deep learning is a subset of machine learning where artificial neural networks, algorithms inspired by the human brain, learn from large amounts of data. Deep learning algorithms are capable of learning to represent the world as a nested hierarchy of concepts, with each concept defined in relation to simpler concepts, and more abstract representations computed in terms of less abstract ones. The skilled reader is referred to for example University of Illinois at Urbana-Champaign; "Al predicts enzyme function better than leading tools." ScienceDaily. ScienceDaily, 30 March 2023.

[0135] <www.scienceda ily.com / releases / 2023 / 03 / 23033017212 l.htm>.

[0136] 2. Contrastive Learning: This is a type of unsupervised learning approach that trains models to learn similar features from similar data points and different features from different data points. An Al tool named 'CLEAN' was recently reported to use this algorithm to predict enzyme function, cf. Gupta, R., Srivastava, D., Sahu, M. et al. Artificial intelligence to deep learning: machine intelligence approach for drug discovery. Mol Divers 25, 1315-1360 (2021) for more details.

[0137] 3. Artificial Neural Networks (ANNs): ANNs are computing systems vaguely inspired by the biological neural networks that constitute animal brains. An ANN is based on a collection of connected units or nodes called artificial neurons, which loosely model the neurons in a biological brain.

[0138] 4. Support Vector Machines (SVMs): SVMs are supervised learning models with associated learning algorithms that analyze data for classification and regression analysis.

[0139] 5. Generative Adversarial Networks (GANs): GANs are a class of artificial intelligence algorithms used in unsupervised machine learning, implemented by a system of two neural networks contesting with each other in a zero-sum game framework.

[0140] These algorithms can be used individually or in combination, depending on the specific requirements of the type of data. The invention according to this aspect can be implemented by means of hardware, software, firmware or any combination of these. The invention or some of the features thereof can also be implemented as software running on one or more data processors and / or digital signal processors.

[0141] The individual elements of an embodiment of the invention may be physically, functionally and logically implemented in any suitable way such as in a single unit, in a plurality of units or as part of separate functional units. The invention may be implemented in a single unit or be both physically and functionally distributed between different units and processors.

[0142] A method for determining a need for adjustment of parenteral supplementation

[0143] Acid / base components (NH4+, HCOs", titratable acids (TA) and pH) have been measured in urine samples from patients with intestinal insufficiency or intestinal failure and receiving parenteral supplementation. Based on the levels of the acid / base components, net acid excretion was determined / calculated (NAE = NH4+excretion + TA excretion - HCOs" excretion). When comparing NAE of short bowel patients with NAE from healthy controls striking differences were observed (see example 6). Such differences can be used to determine the need for adjustment of parenteral supplementation in short bowel patients.

[0144] We note that the methods as disclosed herein are in vitro methods (non-invasive) and are not performed on the human body but on urine samples from patients that have intestinal insufficiency and that have previously received parenteral supplementation. Hence, the parenteral supplementation is administered to the patients and concluded before the urine samples are collected.

[0145] Thus, one aspect of the invention relates to a method for determining a need for adjustment of parenteral supplementation in a subject receiving parenteral supplementation and having intestinal insufficiency, such as short bowel, the method comprising providing a urine sample from said subject; measuring levels of at least three biomarkers in said urine sample;

[0146] - determining / calculating a net acid excretion (NAE) value based on the levels of the at least three biomarkers;

[0147] - comparing the NAE value with a predetermined reference range having an upper and a lower limit; wherein, if the NAE value exceeds the upper limit or if the NAE value is below the lower limit, it is indicative of said subject needing adjustment of parenteral supplementation, or wherein, if the NAE value is within the predetermined reference range, it is indicative of said subject not needing adjustment of parenteral supplementation.

[0148] A further aspect of the present invention relates to a method for determining a need for adjustment of parenteral supplementation in a subject receiving parenteral supplementation, the method comprising providing a urine sample from said subject; measuring levels of at least three biomarkers in said urine sample;

[0149] - determining / calculating a net acid excretion (NAE) value based on the levels of the at least three biomarkers;

[0150] - comparing the NAE value with a predetermined reference range having an upper and a lower limit; wherein, if the NAE value exceeds the upper limit or if the NAE value is below the lower limit, it is indicative of said subject needing adjustment of parenteral supplementation, or wherein, if the NAE value is within the predetermined reference range, it is indicative of said subject not needing adjustment of parenteral supplementation.

[0151] If said subject is considered to have a need of adjustment of parenteral supplementation, one or more components of parenteral supplementation is increased and / or decreased and / or one or more compounds are administered to said subject.

[0152] Hence, by determining and calculating NAE in a urine sample from a subject receiving parenteral supplementation and having intestinal insufficiency, such as short bowel it is possible to accurately perform dynamic adjustment of the parenteral supplementation in the subject and thereby avoid / prevent acid-base imbalance or reestablish systemic acid-base parameters within the normal physiological range.

[0153] In an embodiment of the invention, said compounds are selected from the group consisting of: proton pump inhibitors to reduce production of stomach acid by glands in the lining of the stomach, - acid binders or proton chelators, such as potassium binders, to remove or reduce the amount of acid in the gastrointestinal tract, thereby preventing intestinal absorption of acids, and / or minerals, such as chloride, magnesium, potassium and / or sodium.

[0154] In an embodiment, the adjustment of parenteral supplementation comprises adjusting the amino acid levels in said parenteral supplementation.

[0155] In an embodiment, the adjustment of parenteral supplementation comprises adjusting the net acid / base load in said parenteral supplementation.

[0156] A method for monitoring risk of developing acid-base imbalance

[0157] Based on the levels of the acid / base components, net acid excretion was calculated (NAE = NH4+excretion + TA excretion - HCOs" excretion). When comparing NAE of short bowel patients with NAE from healthy controls striking differences were observed. Such differences can be used to monitor risk of developing acid-base imbalance, such as acidosis or alkalosis, in a subject receiving parenteral supplementation.

[0158] A second aspect of the present invention relates to a method for monitoring risk of developing acid-base imbalance in a subject receiving parenteral supplementation and having intestinal insufficiency, such as short bowel, the method comprising providing a urine sample from said subject; measuring levels of at least three biomarkers in said urine sample;

[0159] - determining / calculating a net acid excretion (NAE) value based on the levels of the at least three biomarkers;

[0160] - comparing the NAE value with a predetermined reference range having an upper and a lower limit; wherein, if the NAE value exceeds the upper limit, it is indicative of said subject being at risk of developing acidic imbalance, or wherein, if the NAE value is below the lower limit, it is indicative of said subject being at risk of developing alkaline imbalance, or wherein, if the NAE value is within the predetermined reference range, it is indicative of said subject not being at risk of developing acid-base imbalance.

[0161] A further aspect of the present invention relates to a method for monitoring risk of developing acid-base imbalance in a subject receiving parenteral supplementation, the method comprising providing a urine sample from said subject; measuring levels of at least three biomarkers in said urine sample;

[0162] - determining / calculating a net acid excretion (NAE) value based on the levels of the at least three biomarkers;

[0163] - comparing the NAE value with a predetermined reference range having an upper and a lower limit; wherein, if the NAE value exceeds the upper limit, it is indicative of said subject being at risk of developing acidic imbalance, or wherein, if the NAE value is below the lower limit, it is indicative of said subject being at risk of developing alkaline imbalance, or wherein, if the NAE value is within the predetermined reference range, it is indicative of said subject not being at risk of developing acid-base imbalance.

[0164] If said subject is considered to be at risk of developing acid-base imbalance, one or more components of a parenteral supplementation are adjusted, or one or more compounds are administered to said subject to decrease or eliminate the risk of developing acid-base imbalance, such as acidosis or alkalosis and thereby reestablish the acid / base balance in said subject to fall within normal physiological ranges. In an embodiment of the invention, said compounds are selected from the group consisting of: proton pump inhibitors to reduce production of stomach acid by glands in the lining of the stomach,

[0165] - acid binders or proton chelators, such as potassium binders, to remove or reduce the amount of acid in the gastrointestinal tract, thereby preventing intestinal absorption of acids, and / or minerals, such as chloride, magnesium, potassium and / or sodium.

[0166] In an embodiment, the acid-base imbalance is acidosis or alkalosis.

[0167] A method for monitoring development of acid-base imbalance

[0168] Based on the levels of the acid / base components, net acid excretion was calculated (NAE = NH4+excretion + TA excretion - HCOs" excretion). When comparing NAE of short bowel patients with NAE from healthy controls striking differences were observed. Such differences can be used to monitor development of acid-base imbalance, such as acidosis or alkalosis, in a subject receiving parenteral supplementation.

[0169] Yet a third aspect of the present invention is to provide a method for monitoring development of acid-base imbalance in a subject receiving parenteral supplementation and having intestinal insufficiency, such as short bowel, the method comprising providing a first urine sample from said subject before said subject receives adjustment of parenteral supplementation;

[0170] - adjusting the parenteral supplementation after said first urine sample has been provided; providing a second urine sample wherein the second urine sample has been obtained after the subject has received the adjusted parenteral supplementation; measuring levels of at least three biomarkers in the first and the second urine sample;

[0171] - determining / calculating a net acid excretion (NAE) based on the levels of the at least three biomarkers in the first and the second urine sample;

[0172] - comparing the NAE value with a predetermined reference range having an upper and a lower limit in the first and the second urine sample; wherein, the difference between the NAE value in the first or the second urine sample exceeding the upper limit and the upper limit is indicative of a degree of acidic imbalance in the subject, or wherein, the difference between the NAE value in the first or the second urine sample below the lower limit and the lower limit is indicative of a degree of alkaline imbalance in the subject, or wherein, if the NAE value in the first or the second urine sample is within the predetermined reference range, it is indicative of said subject not having acidbase imbalance.

[0173] A further aspect of the present invention relates to a method for monitoring development of acid-base imbalance in a subject receiving parenteral supplementation, the method comprising providing a first urine sample from said subject before said subject receives adjustment of parenteral supplementation;

[0174] - adjusting the parenteral supplementation after said first urine sample has been provided; providing a second urine sample wherein the second urine sample has been obtained after the subject has received the adjusted parenteral supplementation; measuring levels of at least three biomarkers in the first and the second urine sample;

[0175] - determining / calculating a net acid excretion (NAE) based on the levels of the at least three biomarkers in the first and the second urine sample;

[0176] - comparing the NAE value with a predetermined reference range having an upper and a lower limit in the first and the second urine sample; wherein, the difference between the NAE value in the first or the second urine sample exceeding the upper limit and the upper limit is indicative of a degree of acidic imbalance in the subject, or wherein, the difference between the NAE value in the first or the second urine sample below the lower limit and the lower limit is indicative of a degree of alkaline imbalance in the subject, or wherein, if the NAE value in the first or the second urine sample is within the predetermined reference range, it is indicative of said subject not having acidbase imbalance.

[0177] If said subject is considered to have a degree of acid-base imbalance, such as acidosis or alkalosis, one or more components of a parenteral supplementation is adjusted, or one or more compounds are administered according to the actual degree of acid-base imbalance, such as acidosis or alkalosis in said subject. The aim is to cure or alleviate acid-base imbalance, such as acidosis or alkalosis and thereby reestablish the acid / base balance in said subject to fall within normal physiological ranges.

[0178] In an embodiment of the invention, said compounds may be selected from the group consisting of: proton pump inhibitors to reduce production of stomach acid by glands in the lining of the stomach,

[0179] - acid binders or proton chelators, such as potassium binders, to remove or reduce the amount of acid in the gastrointestinal tract, thereby preventing intestinal absorption of acids, and / or minerals, such as chloride, magnesium, potassium and / or sodium.

[0180] In a preferred embodiment, the first urine sample is provided 12-24 hours after the subject has received parenteral supplementation and the second urine sample is provided 12-24 hours after the subject has received the adjusted parenteral supplementation.

[0181] In an embodiment, the acid-base imbalance is acidosis or alkalosis.

[0182] The following embodiments apply to all of the above aspects relating to a method.

[0183] In an embodiment, the adjustment of parenteral supplementation comprises adjusting the amino acid levels in said parenteral supplementation.

[0184] In an embodiment, the adjustment of parenteral supplementation comprises adjusting the net acid / base load in said parenteral supplementation.

[0185] In an embodiment, the acidic imbalance is subclinical acidosis, overt acidosis or acidosis.

[0186] In an embodiment, the alkaline imbalance is subclinical alkalosis, overt alkalosis or alkalosis. In an embodiment, the administration is intravenous administration and / or oral administration.

[0187] Parental supplementation and adjustment of parental supplementation may be administered intravenously for administration around the gastrointestinal tract, as patients suffer from impaired or non-functional intestines, resulting in their inability or impaired ability to naturally absorb nutrients and electrolytes.

[0188] In other cases, adjustments of parental supplementation may be administered orally in patients with partially functional intestines. Studies have been conducted showing that administration of glucagon-like peptide-2 (GLP-2) in patients with intestinal insufficiency results in an increased intestinal microvillus length, hence increasing digestion and absorption. Thus, patients not having complete intestinal failure may still be able to absorb nutrients and electrolytes from oral administration.

[0189] In a preferred embodiment, the biomarkers are acid-base components.

[0190] In an more preferred embodiment, the adjustment of parenteral supplementation comprises administration of one or more organic ions such as acetate, lactate, malate, and / or chloride, or one or more amino acids, such as cysteine, methionine, lysin, arginine, histidine, glutamic acid, and / or aspartic acid. Cysteine, methionine, lysin, arginine, and histidine are converted into acids when metabolized. Glutamic acid, and / or aspartic acid are converted into bases when metabolized.

[0191] In a preferred embodiment, the acid components are selected from the group consisting of one or more of the amino acids cysteine, methionine, lysine, arginine and / or histidine.

[0192] In a preferred embodiment, the base components are selected from the group consisting of one or more of the amino acids glutamic acid and / or aspartic acid.

[0193] In an embodiment, the acid-base components are selected from the group consisting of NH4+, HCOs", titratable acids (TA), citrate or pH. As seen in the Example, NH4+, HCOs’, titratable acids (TA), citrate and / or pH were measured.

[0194] In a preferred embodiment, the acid-base components are selected from the group consisting of NH4+, HCOs’ and pH.

[0195] In a more preferred embodiment, the acid-base components are selected from the group consisting of NH4+, titratable acids (TA) and pH.

[0196] In an even more preferred embodiment, the acid-base components are selected from the group consisting of HCOs’, titratable acids (TA) and pH.

[0197] In a most preferred embodiment, the acid-base components are selected from the group consisting of NH4+, HCOs’ and titratable acids (TA).

[0198] In an embodiment, the net acid excretion (NAE) is determined by measuring the level NH4+, HCOS’, and titratable acids (TA) and calculating the NAE using the formula:

[0199] NAE = NH4+excretion + TA excretion - HCOs’ excretion

[0200] Example 2 shows that the NAE formula of the embodiment was successfully used to determine total acid load in short bowel patients.

[0201] Example 6 shows that the NAE can be used to determine the reference range i.e. the predetermined reference range.

[0202] In an embodiment, the intestinal insufficiency is intestinal failure, such as partial or complete intestinal failure.

[0203] In an embodiment, the intestinal failure is short bowel, such as acute phase intestinal failure and / or short bowel.

[0204] A subject having intestinal failure and / or short bowel will require long-term parenteral supplementation. Therefore, the method for determining a need for adjustment of parenteral supplementation and / or monitoring the risk of developing acid-base imbalance must be continuously performed or performed regularly during the period where the subject is receiving parenteral supplementation and having intestinal failure such as short bowel. This is to avoid or prevent acid-base imbalance in the subject by timely and dynamically adjusting the parenteral supplementation depending on the measured NAE value. In that way, the subject will constantly maintain systemic acid-base parameters within a normal physiological range.

[0205] In an embodiment, the short bowel is classified as end-jejunostomy (SB-J), jejunocolonic anastomosis with colon in continuity (SB-JC), or jejunoileocolonic anastomosis with colon in continuity (SB-JIC).

[0206] As can be seen in Example 1, the short bowel patients are classified into SB-J, SB- JC, and SB-JIC.

[0207] In an embodiment, the subject is a mammal, preferably a human.

[0208] In an embodiment, the NAE is urine NAE.

[0209] It is surprisingly shown in the Examples, that urinary NAE is a helpful parameter for differentiated acid-base-assessment in intestinal failure patients. This is due to the fact that urine NAE accounts for renal compensation of acid / base loss.

[0210] In an embodiment, the urine sample is selected from the group consisting of a 24- hour urine sample, an 8-hour urine sample, or a spot urine sample.

[0211] The type of urine sample can be any type of urine sample. The urine sample is preferably a spot urine sample, such as more preferably an 8-hour urine sample or even more preferably a 24-hour urine sample.

[0212] In an embodiment, the upper limit of the NAE value of the predetermined reference range is 80 to 110 mmol per day, preferably 90 to 100 mmol per day, more preferably 94 to 96 mmol per day, most preferably 95 mmol per day.

[0213] The upper limit may be the higher 95% percentile of controls and the lower limit may be the lower 5% percentile of controls, and thereby obtaining a 95% prediction interval.

[0214] In an embodiment, the lower limit of the NAE value of the predetermined reference range is -10 to 5 mmol per day, preferably -5 to 0 mmol per day, more preferably -4 to -3 mmol per day. The upper limit may be the higher 95% percentile of controls and the lower limit may be the lower 5% percentile of controls, and thereby obtaining a 95% prediction interval.

[0215] In an embodiment of the invention, the reference range is an interval having an upper and lower limit being selected from about 96% percentile of controls and about 4% percentile of controls or selected from about 94% percentile of controls and about 6% percentile of controls.

[0216] Use of a net acid excretion (NAE) value

[0217] A fourth aspect of the present invention is to provide use of a net acid excretion (NAE) value for determining a need for adjustment of parenteral supplementation in a subject receiving parenteral supplementation and having intestinal insufficiency, such as short bowel.

[0218] A further aspect of the present invention relates to use of a net acid excretion (NAE) value for determining a need for adjustment of parenteral supplementation in a subject receiving parenteral supplementation.

[0219] In an embodiment, the adjustment of parenteral supplementation comprises adjusting the amino acid levels in said parenteral supplementation.

[0220] In an embodiment, the adjustment of parenteral supplementation comprises adjusting the net acid / base load in said parenteral supplementation.

[0221] A fifth aspect of the present invention is to provide use of a net acid excretion (NAE) value for monitoring risk of developing acid-base imbalance in a subject receiving parenteral supplementation and having intestinal insufficiency, such as short bowel.

[0222] A further aspect of the present invention relates to use of a net acid excretion (NAE) value for monitoring risk of developing acid-base imbalance in a subject receiving parenteral supplementation.

[0223] A sixth aspect of the present invention is to provide use of a net acid excretion (NAE) value for monitoring development of acid-base imbalance in a subject receiving parenteral supplementation and having intestinal insufficiency, such as short bowel.

[0224] A further aspect of the present invention relates to use of a net acid excretion (NAE) value for monitoring development of acid-base imbalance in a subject receiving parenteral supplementation.

[0225] In an embodiment, the net acid excretion (NAE) is determined by measuring the level NH4+, HCOS’, and titratable acids (TA) and calculating the NAE using the formula:

[0226] NAE = NH4+excretion + titratable acids (TA) excretion - HCOs’ excretion

[0227] Example 2 shows that the NAE formula of the embodiment was successfully used to determine total acid load in short bowel patients.

[0228] In an embodiment, the adjustment of the net acid / base load in said parenteral supplementation comprises adjusting acid-base components.

[0229] In a further embodiment, the adjustment of the net acid / base load in said parenteral supplementation comprises adjusting acid-base component, wherein said acid-base components are selected from the group consisting of NH4+, HCOs’, titratable acids (TA), citrate or pH, preferably the acid-base components are selected from the group consisting of NH4+, HCOs’ and titratable acids (TA).

[0230] In an embodiment, the adjustment of parenteral supplementation comprises administration of one or more organic ions such as acetate, lactate, malate, and / or chloride, or one or more amino acids, such as cysteine, methionine, lysin, arginine, histidine, glutamic acid, and / or aspartic acid. Cysteine, methionine, lysin, arginine, and histidine are converted into acids when metabolized. Glutamic acid, and / or aspartic acid are converted into bases when metabolized.

[0231] In an embodiment, the acid-base components are selected from the group consisting of cysteine, methionine, lysine, arginine, histidine, glutamic acid and aspartic acid. In a preferred embodiment, the acid components are selected from the group consisting of one or more of the amino acids cysteine, methionine, lysine, arginine and / or histidine.

[0232] In a preferred embodiment, the base components are selected from the group consisting of one or more of the amino acids glutamic acid and / or aspartic acid.

[0233] In an embodiment, the administration is intravenous administration and / or oral administration.

[0234] Parental supplementation and adjustment of parental supplementation may be administered intravenously for administration around the gastrointestinal tract, as patients suffer from impaired or non-functional intestines, resulting in their inability or impaired ability to naturally absorb nutrients and electrolytes.

[0235] In other cases, adjustments of parental supplementation may be administered orally in patients with partially functional intestines. Studies have been conducted showing that administration of glucagon-like peptide-2 (GLP-2) in patients with intestinal insufficiency results in an increased intestinal microvillus length, hence increasing digestion and absorption. Thus, patients not having complete intestinal failure may still be able to absorb nutrients and electrolytes from oral administration.

[0236] In an embodiment, the acid-base imbalance is acidosis or alkalosis.

[0237] In an embodiment, the acidosis is subclinical acidosis or overt acidosis. In an embodiment, the alkalosis is subclinical alkalosis or overt alkalosis.

[0238] In an embodiment, the intestinal insufficiency is intestinal failure, such as partial or complete intestinal failure.

[0239] In an embodiment, the intestinal failure is short bowel, such as acute phase intestinal failure and / or short bowel.

[0240] In an embodiment, the short bowel is classified as end-jejunostomy (SB-J), jejunocolonic anastomosis with colon in continuity (SB-JC), or jejunoileocolonic anastomosis with colon in continuity (SB-JIC). As can be seen in Example 1, the short bowel patients are classified into SB-J, SB- JC, and SB-JIC.

[0241] In an embodiment, the subject is mammal, preferably human.

[0242] In an embodiment, the NAE is urine NAE

[0243] As can be seen in the Examples, urinary NAE is a helpful parameter for differentiated acid-base-assessment in intestinal failure patients. This is due to the fact that urine NAE accounts for renal compensation of acid / base loss.

[0244] In an embodiment, the urine NAE is obtained from a 24-hour urine sample, an 8- hour urine sample, or a spot urine sample. The type of urine sample can be any type of urine sample. The urine sample is preferably a spot urine sample, such as more preferably an 8-hour urine sample or even more preferably a 24-hour urine sample.

[0245] Further aspects

[0246] A further aspect of the invention related to an in vitro method of treating acidbase imbalance in a subject receiving parenteral supplementation and having intestinal insufficiency, such as short bowel, the method comprising providing a urine sample from said subject; measuring levels of at least three biomarkers in said urine sample;

[0247] - determining / calculating a net acid excretion (NAE) value based on the levels of the at least three biomarkers;

[0248] - comparing the NAE value with a predetermined reference range having an upper and a lower limit; wherein, if the NAE value exceeds the upper limit or if the NAE value is below the lower limit, it is indicative of said subject having acid-base imbalance / disturbances and needing adjustment of parenteral supplementation, or wherein, if the NAE value is within the predetermined reference range, it is indicative of said subject not having acid-base imbalance / disturbances and / or not needing adjustment of parenteral supplementation, wherein if said subject is considered to have acid-base imbalance and / or needing adjustment of parenteral supplementation, one or more components of parenteral supplementation is increased and / or decreased and / or one or more compounds are administered to said subject.

[0249] Hence, by determining and calculating NAE in a urine sample from a subject receiving parenteral supplementation and having intestinal insufficiency, such as short bowel it is possible to accurately perform dynamic adjustment of the parenteral supplementation in the subject and thereby avoid / prevent acid-base imbalance or reestablish systemic acid-base parameters within the normal physiological range.

[0250] Said compounds may be selected from the group consisting of: proton pump inhibitors to reduce production of stomach acid by glands in the lining of the stomach,

[0251] - acid binders or proton chelators, such as potassium binders, to remove or reduce the amount of acid in the gastrointestinal tract, thereby preventing intestinal absorption of acids, and / or minerals, such as chloride, magnesium, potassium and / or sodium.

[0252] Yet another aspect of the present invention relates to an in vitro method of treating acid-base imbalance in a subject receiving parenteral supplementation and having intestinal insufficiency, such as short bowel, the method comprising providing a urine sample from said subject; measuring levels of at least three biomarkers in said urine sample;

[0253] - determining / calculating a net acid excretion (NAE) value based on the levels of the at least three biomarkers;

[0254] - comparing the NAE value with a predetermined reference range having an upper and a lower limit; wherein, if the NAE value exceeds the upper limit, it is indicative of said subject having acidic imbalance, or wherein, if the NAE value is below the lower limit, it is indicative of said subject having alkaline imbalance, or wherein, if the NAE value is within the predetermined reference range, it is indicative of said subject not having acid-base imbalance, wherein if said subject has acid-base imbalance, one or more components of a parenteral supplementation is adjusted, and / or one or more compounds are administered to said subject.

[0255] Said compounds may be selected from the group consisting of: proton pump inhibitors to reduce production of stomach acid by glands in the lining of the stomach,

[0256] - acid binders or proton chelators, such as potassium binders, to remove or reduce the amount of acid in the gastrointestinal tract, thereby preventing intestinal absorption of acids, and / or minerals, such as chloride, magnesium, potassium and / or sodium.

[0257] Another aspect of the invention relates to use of a net acid excretion (NAE) value for treating a subject receiving parenteral supplementation and having intestinal insufficiency, such as short bowel.

[0258] It should be noted that embodiments and features described in the context of one of the aspects of the present invention also apply to the other aspects of the invention. Thus, for example the embodiments / claims relating to the method of the invention also apply to the embodiments relating to the use- embodiments / claims. Hence, individual features mentioned in different claims, may possibly be advantageously combined, and the mentioning of these features in different claims does not exclude that a combination of features is not possible and advantageous.

[0259] Although the present invention has been described in connection with the specified embodiments, it should not be construed as being in any way limited to the presented examples. The scope of the present invention is to be interpreted in the light of the accompanying claim set. In the context of the claims, the terms "comprising" or "comprises" do not exclude other possible elements or steps. Also, the mentioning of references such as "a" or "an" etc. should not be construed as excluding a plurality. All patent and non-patent references cited in the present application, are hereby incorporated by reference in their entirety.

[0260] The invention will now be described in further details in the following non-limiting examples.

[0261] Examples

[0262] Example 1 - Characterization of short bowel cohort Aim

[0263] The aim of this study is to characterize the patient cohort having short bowel (SB).

[0264] Materials and methods

[0265] Patient acquisition

[0266] 24h-urine samples from patients managed by the short bowel outpatient clinic of the University Medical Center Rostock between November 2021 and July 2022 were analyzed. Patient characteristics, as suggested by ESPEN1, were recorded prospectively in a customized database (Microsoft Access, 2010). The study (German Clinical Trials Register: DRKS00021085) was approved by the local ethics committee according to German law (A 2018-0128). Patients older than 18 years with chronic intestinal failure or short-term regained oral autonomy after intestinal failure were included. The information was obtained on remaining small bowel length either from the surgical reports if conclusive, or by subtracting the length of the resected bowel from the presumed initial length of 350 cm. Colon length was expressed as a percentage.

[0267] Patenteral supplementation

[0268] All patients received treatment according to ESPEN guidelines. The composition of parenteral support (PS) from pharmacy prescriptions prior to urine sampling. The prescriptions contained amounts of volume, macronutrients (amino acid, glucose, lipid), electrolytes (sodium, potassium, calcium, magnesia, phosphate, acetate / chloride-ratio), and trace elements from either standard or compounded nutrition bags. If the absolute amount of chloride and acetate was not available, the amounts were calculated by dividing the sum of the associated cations, i.e., sodium and potassium, according to the prescribed acetate / chloride ratio. The potential metabolic acid load as well as metabolizable anions from the amino acid solution were not included in the calculation. The daily composition of PS was calculated as the total weekly supplementation divided by seven.

[0269] Results

[0270] Table 1 shows the demographic, anatomical and clinical characteristics of our patient cohort.

[0271] For further results, Table 3 (Example 7) shows the acid-base parameters identified in short bowel / intestinal insufficiency subjects as a consequence of the various surgical resection types.

[0272] Table 1: Patient characteristics of patient cohort. Means or numbers (n) are shown with standard deviation (SD) or percentage of group respectively. P-values investigating the overall effect of anatomy type on a given parameter were derived from one-way ANOVA tests for continuous parameters and from postestimation Wald tests after logistic regression analyses for binary variables.

[0273] Two patients without intestinal surgery but mucosal disease were allocated as SB- JIC.

[0274] No differences were found between types of SB-anatomy regarding age, sex, body mass index, eGFR, or parenteral energy and volume support.

[0275] Short bowel is classified as end-jejunostomy (SB-J), jejunocolonic anastomosis with colon in continuity (SB-JC), or jejunoileocolonic anastomosis with colon in continuity (SB-JIC). SB-JIC patients had significantly longer residual small intestine than SB-J and SB-JC. Residual small intestine were not significantly different SB-J and SB-JC. SB-JIC had significantly longer colon in continuity than SB-JC. Figure 7 illustrates the different SB classifications. Patients with SB-J received numerical higher volumes of daily PS compared to other types, though this was not statistically significant.

[0276] Conclusion

[0277] The short bowel patient cohort comprises 46 patients (38 on parenteral support and 8 with oral autonomy), which are all characterized in Table 1.

[0278] Example 2 - Urine and blood analysis of the short bowel cohort

[0279] Aim

[0280] Urine and blood samples were collected from the short bowel cohort to determine the venous base excess, anion gap, and acid-base measurements. The parameters were assessed for the whole cohort and categorized by anatomical type. Materials and methods

[0281] Patient cohort

[0282] The patient cohort is characterized in Example 1.

[0283] Data was obtained from 67 time points (visits) in the outpatient clinic from 46 patients. 24h urine collections for urinary acid-base measurements were obtained at 55 visits from 38 patients (21 SB-J; 9 SB-JC; 8 SB-JIC). Seven samples were excluded from urine analysis due to acid-base patterns of unusually high ammonium and bicarbonate concentrations with a concurrently high urine-pH. This pattern indicates contamination with urease-producing bacteria which was confirmed by detection of bacterial species with probable or at least possible ability to produce ammonium by mass spectroscopy in these samples (data not shown)

[0284] Blood and urine tests

[0285] All routine blood and urine tests were performed by the central laboratory of University Medical Center Rostock by routine methods well known by the person skilled in the art. Analyzed blood parameters included hemoglobin, serum electrolytes, venous blood gas analysis, creatinine, urea, hormones (renin, aldosterone, copeptin), albumin and C-reactive protein. Estimated glomerular filtration rate (eGFR) was determined with CKD-EPI equation based on creatinine measurements. Serum anion gap was calculated as Na++ K+- Cl- - HCOs" + 0.25 * (40 - Albumin).

[0286] Routine urine measurements of 24-hour urine collection included electrolytes and creatinine. Collection volume was reported by patients. Said routine urine measurements were prepared by methods well known by the person skilled in the art.

[0287] Urine samples were frozen at -20°C immediately in the outpatient clinic and kept for acid-base analyses. Here, pH, bicarbonate (HCO3 ), titratable acidity (TA), and ammonium (NH4+) were measured as previously described.

[0288] Urine pH

[0289] Urine pH was measured with a pH electrode (Metrohm, Glostrup, Denmark). Urine HCOs" concentration

[0290] Urine HCOs" concentration was measured as total CO2 by measuring the amount of CO2 released from small volume (30 pL) samples injected into IM HCI (3 mL) in an 8 mL funnel-shaped container. A continuous N2 flow (40 mL / min) through the HCI filled container carried the released CO2 through a silicone tubing to a LI-COR LI-7000 CO2analyzer (Lincoln, Nebraska, US). Before entering the CO2 analyzer, the gas was dried by condensation by cooling and passed through a CaCOs containing drying tube. Signal output was processed by LI-7000 software. Sample injection resulted in a transient signal increase lasting up to 200 seconds. The area under the curve was calculated and used to measure of HCOs" content. The concentration of HCOs" was back-calculated from linear standard curves based on known NaHCOs standards (0, 1, 5, 25, 50, and 100 mM). A new standard curve was generated at every system start-up. To control for drifting, a 25 mM NaHCOs standard was measured at the end of every measuring series.

[0291] Urine NH4+concentration

[0292] Urine NH4+concentration was measured using OrionTM High-Performance Ammonia Ion-Selective Electrode (Thermo Scientific, Waltham, USA Cat. No. 9512HPBNWP) with the use of Ammonia pH-adjusting Ionic Strength Adjuster (Thermo Scientific, Waltham, USA Cat. No. 951 211). Urine TA was measured by titration to pH 7.4 after addition of hydrochloric acid. The protocol was based on the method of Chan, J.C.M. (1972) (The rapid determination of urinary titratable acid and ammonium and evaluation of freezing as a method of preservation. Clin. Biochem. 5, 94-98). In short, an equal volume of 0.1 mM hydrochloric acid and sample was mixed and subsequently briefly boiled to eliminate any remaining bicarbonate and carbon dioxide from the sample. Titration was performed with the use of an automatic titrator (Eco-titrator, Metrohm, Glostrup, Denmark).

[0293] Net acid excretion (NAE)

[0294] Net acid excretion (NAE) was calculated as TA + NH4+- HCOs". Total acid load was estimated as NAE + the amount of base infused.

[0295] Statistical analysis

[0296] All patients who provided enough urine volume from 24h collections for acid-base measurements were included in the analysis. 16 patients delivered urine samples on more than one visit. To include all available data while accounting for within-patient errors, mixed effects models for repeated measures were used to estimate differences between SB types and for association analyses. Patient characteristics and PS details were assessed by summary statistics. Specifics for statistical testing are provided with each table and figure. All tests were two-sided and performed at a significance level of 5%. Error-bars represents standard error of the mean unless otherwise specified.

[0297] Results

[0298] Venous base excess, anion gap, and total provision of base equivalents of the entire cohort

[0299] Venous base excess, serum anion gap, and total provision of base equivalents from all samples are shown in Figure 1. Mean venous base excess was 0.6 ± 3.9 mmol / l. 62.5% of all samples were outside the reference interval for venous base excess (-2 to 2 mmol / l). 22% were acidotic and 45% were alkalotic. Mean serum anion gap was 14.1 ± 1.5 mmol / L. 8% were outside the reference interval (8 to 16 mmol / L). SB-JC had a higher mean anion gap than SB-J (+1 mmol / L; 95%-CI 0.1 to 1.8 mmol / L, p<.05). Only 1 / 64 samples were in accordance with anion gap acidosis. Mean parenteral provision of base equivalents was 66 ± 42 mmol / day. Thus, a vast majority (67%) of patients on parenteral support display acid / base imbalance / disturbances.

[0300] Venous base excess and urinary acid base parameters categorized by anatomical type

[0301] Figure 2 shows venous base excess and acid-base measurements from 24h urine collections for each type of SB-anatomy. The following estimates are least-square means followed by 95% Cis derived from a mixed-effects analysis for repeated measures. Venous base excess was significantly lower in SB-JC (-4.1 mmol / L; 95% CI: -6.3 to -1.9 mmol / L; p<.001) and SB-JIC (-3.1 mmol / L; 95% CI: -5.6 to -0.5 mmol / L; p<.05) compared to SB-J (2.5 mmol / L). Urine pH was significantly lower in SB-JC (-0.8; -1.4 to -0.2; pc.01) and SB-JIC (-0.8; -1.4 to -0.1; pc.01) compared to SB-J. None of the groups showed overt metabolic acidosis on average, but 10 out of 19 samples from SB-JC patients (vs. 2 / 34 patients with SB-J) were acidotic (odds ratio (OR) for acidosis: 17.8, p=.001). In contrast, 19 / 34 samples from SB-J patients were alkalotic, while only 3 / 19 SB-JC samples were alkalotic (OR for alkalosis: 6.8, p=0.008)

[0302] Urine measurements revealed higher ammonium excretion (+38.3 mmol / day; 13.5 to 63; p<.01), higher TA excretion (+27.6 mmol / day; 0 to 55.2; p<.05), and lower bicarbonate excretion (-27.6 mmol / day; -46.8 to -8.3; p<.01) in SB-JC compared to SB-J. Thus, the calculated NAE was significantly higher in SB-JC compared to SB-J (+93 mmol / day; 50.7 to 135.4; p<.001). SB-JC had a higher ammonium excretion than SB-JIC (33.8 mmol / day, 0.2 to 67.4, p<.05). Otherwise, no differences were found between SB-JC and SB-JIC. Differences between SB-J and SB-JC were consistent after adjustment for age, sex, eGFR, acetate supplementation and small bowel length (Table 2).

[0303] Table 2: Estimated differences in acid-base parameters between SB-J (type 1 ) and SB-JC (type 2) patients adjusted for: sex, age, GFR, and acetate supplementation. Estimates are least-square means followed by 95% Cis derived from a mixed-effects analysis for repeated measures. Observations: 40 (urine measurements) and 52 (base excess). Individuals: 29 (urine measurements) and

[0304] In this multivariate analysis, only anatomy type, eGFR, and acetate supplementation was significantly associated with NAE.

[0305] Conclusion Venous base excess, serum anion gap, and total provision of base equivalents from all patients in the cohort are shown in Figure 1.

[0306] Urine measurements revealed higher ammonium excretion, higher TA excretion, and lower bicarbonate excretion in SB-JC compared to SB-J. Thus, the calculated NAE was significantly higher in SB-JC compared to SB-J.

[0307] Renal function was only mildly reduced (eGFR 62.5±32.5 ml / min / 1.73m2) and anion gap was within the upper limit of the normal range. Thus, acidotic deviations were not the result of renal insufficiency or an added acid such as lactic acid, ketonic acid or uremic acids but rather reflected intestinal loss of bicarbonate. They were also not the result of chloride infusion acidosis.

[0308] Example 3 - Provision of parenteral base equivalents and its impact on base excess

[0309] Aim

[0310] The aim of the present example is to determine parenteral base equivalents and its impact on base excess.

[0311] Materials and methods

[0312] Patient cohort

[0313] The patient cohort is characterized in Example 1.

[0314] Blood and urine tests

[0315] Blood and urine tests from each patient were obtained and measurements were performed as described in Example 2.

[0316] Results

[0317] Patients with SB-J received numerically larger amounts of acetate (72 ± 41 mmol / day) than SB-JC (58 ± 46 mmol / day) and SB-JIC (58 ± 39 mmol / day), but the differences were not statistically significant (Figure 3A).

[0318] During steady-state conditions, NAE will correspond to the net endogenous acid production. As base supplementation will mask an increased endogenous need for acid excretion, the amount of infused base equivalents was added to NAE to estimate the total acid load. Total acid load was significantly higher in SB-JC patients that in SB-J patients (+78.2 mmol / day; 95%-CI: 38.7 to 117.8; p<.001, figure 3B). After adjustment for sex, age, eGFR, and small bowel length, SB-JC patients still had a markedly higher total acid load and lower BE compared to patients with SB- J (Table 2). Adjusted total acid load was 84.5 mmol / day (95% CI: 41.3 to 127.7) higher and BE was -4 mmol / L (95% CI: -6.3 to -1.9) lower in SB-JC compared to SB-J. Compared with a healthy control cohort, the total acid load was significantly higher in all SB-types but varied from an ~2-fold (SB-J) to an ~5-fold (SB-JC) increased acid load (Figure 3B)

[0319] Total acid load strongly correlated with the measured base excess (Figure 3C). SB-JC patients were predominantly acidotic and were thus under-buffered regarding base provision in the parenteral support, while SB-J patients had a positive base excess and were thus predominantly over-buffered (Figure 3C).

[0320] Conclusion

[0321] Intestinal base loss was significantly higher in SB-JC than in SB-J. SB-JC patients had 84.5 mmol / day higher acid load than SB-J patients. In concordance with this large difference, SB-JC patients had a small but significantly acidic base excess indicating under-buffering, while SB-J patients had a slightly positive base excess, indicating too intense buffering.

[0322] In conclusion, short bowel patients with jejunocolonic anastomosis (SB-JC) have a ~4.4-fold higher total acid load compared to healthy controls due to bicarbonate loss from the mid-to-distal colon. This results in lower base excess and a higher risk for acidic imbalance or even metabolic acidosis in many of these patients, indicating incomplete compensation by the kidney and insufficient provision of base equivalents in their parenteral support. In contrast, SB-J patients may often be over-buffered. To rebalance acid-base homeostasis, base equivalents added to the parenteral support are best titrated individually based on base excess and urine net acid excretion. A reasonable approach wil be to adjust base infusion with the aim of achieving a base excess in the normal range (-2 to +2 mmol / L) while maintaining urine NAE in a near-to-normal range (e.g., -20 to 100 mmol / day). This approach will result in an optimized acid-base balance, thus more stable homeostatic conditions for the patients.

[0323] Example 4 - Role of the residual small intestine for total acid load Aim The aim of the present example is to determine the role of the residual small intestine for total acid load.

[0324] Materials and methods

[0325] Patient cohort

[0326] The patient cohort is characterized in Example 1.

[0327] Blood and urine tests

[0328] Blood and urine tests from each patient were obtained and measurements were performed as described in Example 2.

[0329] Results

[0330] A shorter small bowel was associated with a higher BE in SB-J but not in SB-JC or SB-JIC (Figure 4A-B). Further, a shorter small bowel was associated with a higher amount of base infusion in SB-J, but not in SB-JC or SB-JIC (figure 4C-D). There was no association between total acid load and small bowel length in any SB-type (figure 4E-F). Taken together these data argue against a significant role of the residual small intestine in the pathophysiology of metabolic acidosis in SB- JC and SB-JIC.

[0331] Conclusion

[0332] Small bowel length was not significantly different between SB-J and SB-JC and small bowel length was not related to intestinal base loss in either SB-J nor in SB- JC or SB-JIC. Taken together, this indicates that this base loss was brought about by the presence of colon in continuity specifically mid-distal colon in continuity because it is the right side of the colon that is resected in SB-JC patients.

[0333] Example 5 - Correlation between parenteral support components and acid-base-parameters

[0334] Aim

[0335] The aim of the present example is to determine the association between parenteral support components, such as acetate, chloride, and sodium, and acid- base-parameters, such as net acid excretion (NAE) and base excess (BE).

[0336] Materials and methods Patient cohort

[0337] The patient cohort is characterized in Example 1.

[0338] Blood and urine tests

[0339] Blood and urine tests from each patient were obtained and measurements were performed as described in Example 2.

[0340] Results

[0341] The amount of amino acids, glucose, lipid, energy, or volume per week were not associated with either NAE or BE. Receiving higher amounts of acetate, chloride, and sodium infusion were associated with a lower NAE (persistent after adjustment for BE) and a higher BE (Figure 5 ) and moderately associated with receiving higher amounts of potassium, calcium, and magnesium (not associated after adjustment for BE) (data not shown). Acetate concentration (data not shown) as well as acetate / chloride ratio of parenteral support did not correlate with NAE or BE (Figure 5).

[0342] Conclusion

[0343] Receiving higher amounts of acetate, chloride, and sodium infusion were associated with a lower NAE (persistent after adjustment for BE) and a higher BE. Lower NAE and a higher BE were moderately associated with receiving higher amounts of potassium, calcium, and magnesium, however no association was seen after adjustment for BE.

[0344] Example 6 - Identification of predetermined reference range

[0345] Aim

[0346] The aim of the present example is to identify a reference range for the net acid excretion (NAE) in healthy control individuals which can be used to identify subjects with intestinal failure that is above or is below said reference range.

[0347] Materials and methods

[0348] Patient cohort

[0349] The patient cohort is characterized in Example 1. For this example, all patients are pooled in one group i.e. 46 subjects with intestinal failure are analysed together and their NAE is determined / calculated. Blood and urine tests

[0350] Blood and urine tests from each patient were obtained and measurements were performed as described in Example 2.

[0351] Conclusion

[0352] Figure 6 shows 24h NAE in 46 subjects with intestinal failure and 25 healthy control individuals. The dotted line represents the lower 5% percentile (-3.78 mmol / day) and the higher dotted line represents the 95% percentile (95.18 mmol / day) of the controls. NAE-values within the dotted lines hereby provide a predetermined reference range for normal acid-base status in healthy control individuals. This predetermined reference range can be used to evaluate if subjects falling outside the predetermined reference need adjustment of their parenteral supplementation. Accordingly, the intestinal failure subjects that have NAE-values below the lower dotted line need to excrete to much base and the intestinal failure subjects that have NAE-values above the upper dotted line need to excrete too much acid i.e. said subjects have acid-base disturbances and may be at risk of developing acidosis or alkalosis. Hence, these two groups of subjects would benefit from adjustment of their parenteral supplementations in order to arrive at NAE-values that are within the control / reference range i.e. within normal physiological rage.

[0353] In summary, it is suggested that individual parenteral supplementation must become adjusted so that subjects / patients display full normalization of systemic acid-base parameters and reach urine NAE within the normal physiological rage (i.e. as defined by the predetermined reference range shown above).

[0354] Thus, the inventors have demonstrated an improved in vitro and non-invasive method for determining the risk of having and / or degree of acidosis or alkalosis in intestinal failure patients / subjects / short bowl patients. Also, in particular the inventors have identified a more efficient and / or reliable method for determining the risk of having and / or degree of acidosis or alkalosis in intestinal failure patients / subjects / short bowel patients in order to be able to determine the need of adjustment of parenteral supplementation. For this purpose, they have used urine NAE value. Example 7 - Acid-base parameters as a function of anatomy type

[0355] Aim

[0356] The aim of this study is to characterize the acid-base parameters of subjects in the short bowel patient cohort.

[0357] Results

[0358] Table 3 shows the values identified in short bowel / intestinal insufficiency subjects as a consequence of the various surgical resection types. The data is from the patient cohort, as presented in Example 1.

[0359] Table 3 shows the estimated acid-base parameters for type 1 patients and the difference between type 1, 2, and 3 patients. Estimates are least-square means derived from a mixed-effects analysis for repeated measures. SB-types means are followed by SEMs. Differences between SB types are followed by 95% Cis.

[0360] Observations: 48 (urine measurements) and 64 (base excess). Individuals: 34 (urine measurements) and 44 (base excess). *p<0.05, **p<0.01, ***p<0.001

[0361]

Claims

Claims1. A method for determining a need for adjustment of parenteral supplementation in a subject receiving parenteral supplementation and having short bowel, the method comprising providing a urine sample from said subject; measuring levels of at least three biomarkers in said urine sample;- determining / calculating a net acid excretion (NAE) value based on the levels of the at least three biomarkers;- comparing the NAE value with a reference range having an upper and a lower limit; wherein, if the NAE value exceeds the upper limit or if the NAE value is below the lower limit, it is indicative of said subject needing adjustment of parenteral supplementation, or wherein, if the NAE value is within the reference range, it is indicative of said subject not needing adjustment of parenteral supplementation.

2. A method for monitoring risk of developing acid-base imbalance in a subject receiving parenteral supplementation and having short bowel, the method comprising providing a urine sample from said subject; measuring levels of at least three biomarkers in said urine sample;- determining / calculating a net acid excretion (NAE) value based on the levels of the at least three biomarkers;- comparing the NAE value with a reference range having an upper and a lower limit; wherein, if the NAE value exceeds the upper limit, it is indicative of said subject being at risk of developing acidic imbalance, or wherein, if the NAE value is below the lower limit, it is indicative of said subject being at risk of developing alkaline imbalance, or wherein, if the NAE value is within the reference range, it is indicative of said subject not being at risk of developing acid-base imbalance.

3. A method for monitoring development of acid-base imbalance in a subject receiving parenteral supplementation and having short bowel, the method comprising providing a first urine sample from said subject before said subject receives adjustment of parenteral supplementation;- adjusting the parenteral supplementation after said first urine sample has been provided; providing a second urine sample wherein the second urine sample has been obtained after the subject has received the adjusted parenteral supplementation; measuring levels of at least three biomarkers in the first and the second urine sample;- determining / calculating a net acid excretion (NAE) based on the levels of the at least three biomarkers in the first and the second urine sample;- comparing the NAE value with a reference range having an upper and a lower limit in the first and the second urine sample; wherein, the difference between the NAE value in the first or the second urine sample exceeding the upper limit and the upper limit is indicative of a degree of acidic imbalance in the subject, or wherein, the difference between the NAE value in the first or the second urine sample below the lower limit and the lower limit is indicative of a degree of alkaline imbalance in the subject, or wherein, if the NAE value in the first or the second urine sample is within the reference range, it is indicative of said subject not having acid-base imbalance.

4. The method according to any of claims 2 or 3, wherein the acidic imbalance is subclinical acidosis, overt acidosis or acidosis.

5. The method according to any of claims 2 or 3, wherein the alkaline imbalance is subclinical alkalosis, overt alkalosis or alkalosis.

6. The method according to any of the preceding claims, wherein the net acid excretion (NAE) is determined by measuring the level NH4+, HCOs", and titratable acids (TA) and calculating the NAE using the formula:NAE = NH4+excretion + titratable acids (TA) excretion - HCOs" excretion.

7. The method according to any of the preceding claims, wherein the NAE is urine NAE.

8. The method according to any of the preceding claims, wherein the short bowel is selected from the group consisting of end-jejunostomy (SB-J), jejunocolonic anastomosis with colon in continuity (SB-JC), or jejunoileocolonic anastomosis with colon in continuity (SB-JIC).

9. The method according to any of the preceding claims, wherein the urine sample is selected from the group consisting of a 24-hour urine sample, an 8-hour urine sample, or a spot urine sample.

10. The method according to any of the preceding claims, wherein the upper limit of the NAE value of the reference range is 80 to 110 mmol per day, preferably 90 to 100 mmol per day, more preferably 94 to 96 mmol per day, most preferably 95 mmol per day.

11. The method according to any of the preceding claims, wherein the lower limit of the NAE value of the reference range is -10 to 5 mmol per day, preferably -5 to 0 mmol per day, more preferably -4 to -3 mmol per day.

12. The method according to any of the preceding claims, wherein the reference range is an interval having an upper and lower limit being selected from about 96% percentile of controls and about 4% percentile of controls or selected from about 94% percentile of controls and about 6% percentile of controls.

13. The method according to any of the preceding claims, wherein the biomarkers are acid-base components.

14. The method according to claims 1, 3-13, wherein the adjustment of parenteral supplementation comprises adjusting the amino acid levels in said parenteral supplementation.

15. The method according to claims 1, 3-14, wherein the adjustment of parenteral supplementation comprises adjusting the net acid / base load in said parenteral supplementation.

16. The method according to claims 1, 3-15, wherein the adjustment of parenteral supplementation comprises administration of one or more organic ions such as acetate, lactate, malate, and / or chloride, or one or more amino acids, such as cysteine, methionine, lysin, arginine, histidine, glutamic acid, and / or aspartic acid.

17. The method according to claim 16, wherein the administration is intravenous administration and / or oral administration.

18. The method according to claim 13, wherein the acid-base components are selected from the group consisting NH4+, HCCU’ and titratable acids (TA).

19. The method according to any of the preceding claims, wherein the subject is a mammal, preferably a human.

20. Use of a net acid excretion (NAE) value for determining a need for adjustment of parenteral supplementation in a subject receiving parenteral supplementation and having short bowel.

21. Use of a net acid excretion (NAE) value for monitoring risk of developing acid-base imbalance in a subject receiving parenteral supplementation and having short bowel.

22. Use of a net acid excretion (NAE) value for monitoring development of acidbase imbalance in a subject receiving parenteral supplementation and having short bowel.

23. The use according to any of claims 20-22, wherein the NAE is urine NAE from a urine sample.

24. The use according to claim 23, wherein the urine sample is selected from the group consisting of a 24-hour urine sample, an 8-hour urine sample, or a spot urine sample.

25. The use according to any of claims 20-24, wherein the net acid excretion (NAE) is determined by measuring the level NH4+, HCOs", and titratable acids (TA) and calculating the NAE using the formula:NAE = NH4+excretion + titratable acids (TA) excretion - HCOs" excretion.

26. The use according to any of claims 20, 23-25, wherein the adjustment of parenteral supplementation comprises adjusting acid-base component, wherein said acid-base components are selected from the group consisting of NH4+, HCOs" and titratable acids (TA).

27. The use according to claim 20, 23-26, wherein the adjustment of parenteral supplementation comprises administration of one or more organic ions such as acetate, lactate, malate, and / or chloride, or one or more amino acids, such as cysteine, methionine, lysin, arginine, histidine, glutamic acid, and / or aspartic acid.

28. The use according to claim 20, 23-27, wherein the adjustment of parenteral supplementation comprises adjusting the amino acid levels in said parenteral supplementation.

29. The use according to claim 20, 23-28, wherein the adjustment of parenteral supplementation comprises adjusting the net acid / base load in said parenteral supplementation.

30. The use according to claim 29, wherein the adjustment of the net acid / base load in said parenteral supplementation comprises adjusting acid-base components.

31. The use according to claim 30, wherein, the acid-base components are selected from the group consisting of cysteine, methionine, lysine, arginine, histidine, glutamic acid and aspartic acid.

32. The use according to any of claims 21-25, wherein the acid-base imbalance is acidosis, such as subclinical acidosis or overt acidosis, or alkalosis, such as subclinical alkalosis or overt alkalosis.

33. The use according to any of claims 20-32, wherein the short bowel is selected from the group consisting of end-jejunostomy (SB-J), jejunocolonic anastomosis with colon in continuity (SB-JC), or jejunoileocolonic anastomosis with colon in continuity (SB-JIC).

34. The use according to any of claims 20-33, wherein the subject is a mammal, preferably a human.

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