Preparation method of flavoring agent

Abstract

The present invention relates to a process for the preparation of a flavoring comprising the steps of: (i) providing a reaction mixture comprising a yeast extract, the yeast extract comprises at least 0.5 wt% of a glycosamine and / or a derivative thereof, based on the dry matter weight of the yeast extract, and at least 1.0 wt% of a thiol-containing compound, based on the weight of the dry yeast extract; at least 2% by weight, based on the weight of the reaction mixture, of a mineral acid or organic acid; at least 10% by weight of water, based on the weight of the reaction mixture; (ii) heating the reaction mixture at a temperature of 90 to 160 DEG C; wherein the heated reaction mixture is acidic such that 1% by weight of the diluent in the softened water has a pH of 3.5 to 6 at 20 DEG C; and (iii) optionally drying the product.

Description

[0001] This application is a divisional application of Chinese patent application No. 201980070093.6, filed on October 15, 2019, with the invention name “Method for preparing seasoning”. Technical Field

[0002] The present invention relates to a method for producing a flavoring agent, a flavoring agent obtainable by said method and a food product comprising said flavoring agent. Background Art

[0003] Many consumer food products such as soups, ready meals, bouillon cubes contain flavourings to ensure that the product meets the taste expectations of the consumer. Providing meat flavourings for these food products has received particular attention.

[0004] Usually, meat flavoring is synthetically derived, prepared by the Maillard reaction and the Strecker reaction of sulfur-containing amino acids and sugars. Sulfur-containing amino acids can be provided by free amino acids (e.g., cysteine) or peptides containing sulfur-containing amino acids such as glutathione. The sugar used is usually glucose, ribose, arabinose or xylose. Mottram et al. described in J.Braz.Chem.Soc.9,261 (1998) that cysteine ​​and pure inositol monophosphate (IMP, a ribonucleotide) are combined by heating for 1h at 140 ° C and 0.28MPa (2.7 bar) pressure at pH 3.0, 4.5 and 6. The defect of this chemical synthesis is that according to the EFFA (European Flavor and Fragrance Association) guidelines, the resulting salty and fresh flavoring cannot be described as "natural". Consumers increasingly expect to be labeled as foods containing only natural ingredients. Therefore, it is necessary to provide a preparation method for natural salty and fresh flavoring.

[0005] JP 2003-169627 Example 1 discloses a method for preparing a meat seasoning, wherein 7 parts by weight of a yeast extract containing 5'-nucleotides (produced by Kohjin, 5'-nucleotide content 36%), a yeast extract containing glutathione (yeast extract Aromild U, glutathione content 8%), 2.5 parts by weight of xylose, 7 parts by weight of powdered dextrin (produced by SanwaStarch Co., Ltd., Sandech #70) and 14.5 parts by weight of sodium chloride are added to 50 parts by weight of water and heated at 105° C. for 65 minutes. After the reaction is completed, the reaction solution is dried with a spray dryer to obtain a seasoning. The method of JP 2003-169627 has the disadvantage that pure xylose and dextrin produced by a chemical method are added, so the resulting seasoning cannot be called natural.

[0006] Another disadvantage of the heat-mediated Maillard-based seasoning of JP 2003-169627 is the undesirable cooking or burning smell. The purpose of JP 2016-174587 is to overcome this drawback by including a proline source. Example 5 discloses a composition comprising 49.0 wt% glutathione yeast extract, 4.0 wt% 5'-ribonucleotide-containing yeast extract, 1.4 wt% fructose syrup, 5.0 wt% proline-containing yeast extract and 40.6 wt% water, which is heated at 110°C for 30 minutes.

[0007] There remains a need to provide methods for preparing meat flavorings that meet consumer demands and regulatory requirements. Summary of the invention

[0008] In a first aspect, a method for preparing a flavoring agent is provided, comprising the steps of:

[0009] (i) providing a reaction mixture, the reaction mixture comprising:

[0010] a. a yeast extract, the yeast extract comprising at least 0.5% by weight of sugar amines and / or derivatives thereof, based on the weight of the dry matter of the yeast extract, and at least 1.0% by weight of thiol-containing compounds, based on the weight of the dry yeast extract;

[0011] b. at least 2% by weight of the reaction mixture in an amount of an acid;

[0012] c. at least 10% by weight of water, based on the weight of the reaction mixture;

[0013] (ii) heating the reaction mixture at a temperature of 90-160° C.,

[0014] wherein the heated reaction mixture is acidic so that a 1 wt % dilution in demineralized water has a pH of 3.5 to 6 at 20° C.;

[0015] (iii) optionally drying the product,

[0016] wherein the reaction mixture is heated for a period of (t) hours according to the following formula (I):

[0017] t=4096e -bT (I)

[0018] Among them: 90℃≤T≤160℃, and 0.06≤b≤0.07.

[0019] The inventors of the present invention have surprisingly found that a seasoning with a meaty taste is provided by a process wherein a yeast extract comprising 0.5 wt. % of a sugar amine and / or a derivative thereof and 1 wt. % of a thiol-containing compound, based on the weight of the dry matter of the yeast extract, is heated at a temperature of 90-160° C. in the presence of an acid.

[0020] Surprisingly, the conversion of sulfhydryl-containing compounds and sugar amines and / or derivatives thereof present in the yeast extract proceeds efficiently at / above 90°C to provide a meaty flavour without the formation of competing off-flavours.

[0021] The present invention has the following advantages over the prior art methods: no chemically processed reducing sugars such as glucose-fructose syrup are required in the reaction mixture to provide the meat flavoring. Thus, the method of the present invention meets the consumer demand for natural flavorings. DETAILED DESCRIPTION

[0022] As used herein, the word "comprising" means "including," but does not necessarily mean "consisting of" or "composed of." In other words, the listed steps or options need not be exhaustive.

[0023] Unless otherwise specified, numerical ranges expressed in the format of "x to y" or "xy" are understood to include x and y. When multiple preferred ranges are described in the format of "x to y" or "xy" for a specific feature, it is understood that all ranges combining the different endpoints are also contemplated. For the purposes of the present invention, ambient temperature is defined as a temperature of about 20°C.

[0024] Unless otherwise indicated, weight percents (wt %) are based on the total weight of the composition.

[0025] The term "yeast" as used herein means an organism belonging to the Saccharomycetaceae family, preferably to the genus Saccharomyces. Yeast in the context of the present invention may be any type of yeast.

[0026] The term "yeast extract" as used herein refers to a composition comprising water-soluble components of yeast cells, which are primarily composed of amino acids, peptides, carbohydrates and salts. Yeast extract is produced by hydrolysis (i.e., autolysis or hydrolysis) of peptide bonds by enzymes naturally present in edible yeast or by addition of food-grade enzymes (Food Chemical Codes). In the context of the present invention, yeast extract is synonymous with yeast hydrolysate or yeast autolysate, i.e., a soluble extract from yeast containing hydrolyzed proteins.

[0027] The term "glycosamine and / or derivatives thereof" refers to a sugar group covalently linked to an amino group. The sugar group is formed by removing a hydroxyl group from a hemiacetal functional group of a monosaccharide (e.g., ribose or glucose). The amino group may be provided, for example, by a purine group, a pyrimidine group, or a pyridine group. The derivative may include a phosphorylated hydroxyl group, such as in a 5'-ribonucleotide.

[0028] The term "ribonucleotide" as used herein refers to a mixture of guanosine monophosphate (5'-GMP), cytidine monophosphate (5-CMP), uridine monophosphate (5'-UMP) and further adenosine monophosphate (5'-AMP) and / or inosine monophosphate (5'-IMP), wherein the 5-IMP in the mixture is obtained by partial or complete conversion of 5'-AMP into 5'-IMP. The term "ribonucleotide" as used herein means free 5'-nucleotides or salts thereof. The weight percentage (wt%) of 5'-nucleotides in yeast extracts is based on the salt-free dry matter weight of the composition and is calculated as the disodium salt heptahydrate (2Na.7Aq) of 5'-nucleotides. Salt-free does not mean that the composition of the present invention cannot contain salts, but rather that salts are excluded from the composition in the calculation of the wt%. Ribonucleotides and nucleotides are used interchangeably within the scope of this application.

[0029] As used herein, the term "ribonucleoside" refers to a mixture of guanosine, cytidine monophosphate, uridine, adenosine and / or inosine.

[0030] The amount of ribonucleotides and / or ribonucleosides present in the reaction mixture can be determined by 1 H NMR determination.

[0031] The yeast extract according to the invention comprises on the one hand "total dry matter" and on the other hand "water". The sum of the weights of total dry matter and water is defined as 100%. For example, a yeast extract may comprise 15% by weight of water and 85% by weight of total dry matter.

[0032] The "total dry matter" in a yeast extract comprises on the one hand "yeast dry matter" or "salt-free dry matter" and on the other hand "non-yeast dry matter" such as (added) salt. The "yeast dry matter" or "salt-free dry matter" in a yeast extract may include amino acids, peptides, lipids, polysaccharides, 5'-ribonucleotides, etc.

[0033] Thus, a yeast extract may have the following composition:

[0034] 60% water;

[0035] 40% total dry matter, of which 8% salt (sodium chloride) = 20% by weight of the total dry matter;

[0036] 32% yeast dry matter, which = 80% of the total dry matter;

[0037] 6% 5'-ribonucleotides, which = 18.75% (6 / 32) of yeast dry matter; and

[0038] 26% other components such as amino acids, peptides, lipids, and polysaccharides.

[0039] As used herein, the term "acid" refers to a compound that is a Bronsted acid.

[0040] The term "sulfhydryl-containing compound" as used herein means a compound having an -SH group such as cysteine, and a masked -SH compound such as a disulphide-containing compound (-SS) such as cystine. The total amount of -SH can be determined according to the method in Proteins and Proteomics by Richard J. Simpson (CSHL Press, Cold Spring Harbor, NY, USA, 2003).

[0041] According to the present invention, the reaction mixture comprises yeast extract, acid and water. Preferably, the reaction mixture comprises at least 30% by weight of yeast extract, preferably at least 40% by weight of yeast extract, more preferably at least 50% by weight of yeast extract, even more preferably at least 60% by weight, still more preferably at least 70% by weight of yeast extract, most preferably at least 80% by weight, based on the weight of the reaction mixture. In other words, the reaction mixture comprises 30-80% by weight of yeast extract, preferably the reaction mixture comprises 40-75% by weight of yeast extract, more preferably the reaction mixture comprises 45-65% by weight of yeast extract, based on the weight of the reaction mixture.

[0042] The yeast extract may belong to the family Saccharomyces. In a specific embodiment, the yeast is of the genus Saccharomyces, such as Saccharomyces cerevisiae or Saccharomyces uvarum. In another embodiment, the yeast is of the genus Kluyveromyces, such as Kluyveromyces fragilis. In yet another embodiment, the yeast is of the genus Candida, such as Candida utilis, also known as Turula yeast.

[0043] The reaction mixture defined herein comprises at least 0.5% by weight of a sugar amine and / or its derivatives based on the weight of the dry matter of the yeast extract and at least 1% by weight of a sulfhydryl-containing compound based on the weight of the dry matter of the yeast extract. The sugar amine and / or its derivatives and the sulfhydryl-containing compound may be provided by the same or different yeast extracts. In other words, the sugar amine and / or its derivatives and the sulfhydryl-containing compound may be provided by a first yeast extract and a second yeast extract, respectively, wherein in certain embodiments, the first yeast extract and the second yeast extract may be the same.

[0044] In a preferred embodiment, step (i) comprises providing a first yeast extract and a second yeast extract, wherein the first yeast extract comprises at least 0.5 wt. % of sugar amines and / or derivatives thereof, based on the weight of the dry matter of the yeast extract, and the second yeast extract comprises at least 1 wt. % of thiol-containing compounds, based on the weight of the dry matter of the yeast extract.

[0045] In another preferred embodiment, step (i) comprises providing a yeast extract comprising at least 0.5 wt. % of sugar amines and / or derivatives thereof, based on the weight of the dry matter of the yeast extract, heating the yeast extract in the presence of an acid, and subsequently adding a yeast extract comprising at least 1 wt. % of sulfhydryl-containing compounds, based on the weight of the dry matter of the yeast extract. The resulting reaction mixture is then heated at a temperature of 90-160° C. in the presence of an acid.

[0046] Preferably, the yeast extract added in step (i) comprises at least 1 wt.-%, preferably at least 2 wt.-%, preferably at least 5 wt.-%, more preferably at least 10 wt.-%, even more preferably at least 15 wt.-%, most preferably at least 20 wt.-% of sugar amines and / or derivatives thereof, based on the weight of the dry matter of the yeast extract.

[0047] Typically, the reaction mixture comprises a yeast extract comprising 20-75 wt. %, more preferably 30-60 wt. %, even more preferably 40-50 wt. % of sugar amines and / or derivatives thereof, based on the weight of the reaction mixture.

[0048] The reaction mixture typically comprises a yeast extract comprising 20-75 wt %, more preferably 30-60 wt %, even more preferably 40-50 wt % of the thiol-containing compound based on the weight of the reaction mixture.

[0049] Preferably, the reaction mixture comprises a yeast extract comprising 20-75 wt. %, more preferably 30-60 wt. %, even more preferably 40-50 wt. % of sugar amines and / or derivatives thereof and thiol-containing compounds, based on the weight of the reaction mixture.

[0050] In a preferred embodiment, the sugar amine and / or its derivative contains a ribose moiety. It has been found that using ribose-containing sugar amines according to the method of the present invention, compounds such as 2-methyl-3-furanthiol, 2-methyl-3-(methylthio) furan and bis(2-methyl-3-furanyl) disulfide are formed, and these furan-containing compounds provide a characteristic meaty flavor.

[0051] In certain embodiments, the sugar amine and / or its derivative is a ribonucleotide and / or a ribonucleoside. Preferably, the yeast extract comprises at least 0.5 wt. %, preferably at least 1 wt. %, preferably at least 5 wt. %, more preferably at least 10 wt. %, even more preferably at least 15 wt. %, most preferably at least 20 wt. % ribonucleotides based on the weight of the dry matter of the yeast extract.

[0052] In another preferred embodiment, the sugar amine is a ribonucleotide and / or a ribonucleoside. Without wishing to be bound by theory, the inventors postulate that heating the ribonucleotide in the presence of an acid causes the ribonucleotide and / or ribonucleoside to degrade to release ribose, which can participate in a reaction with the sulfhydryl-containing compound present to generate a flavoring compound.

[0053] The sulfhydryl-containing compound can be a protein, peptide or amino acid containing a -SH group. In other words, the thiol group can exist as a "free amino acid" or as a part of a peptide and / or protein. Preferably, the sulfhydryl-containing compound is a peptide containing glutathione or cysteine ​​(preferably glutathione).

[0054] In a preferred embodiment, the yeast extract comprises at least 2 wt.-%, preferably at least 5 wt.-%, preferably at least 10 wt.-%, more preferably at least 15 wt.-%, even more preferably at least 20 wt.-%, most preferably at least 25 wt.-% of thiol-containing compounds, based on the weight of the dry matter of the yeast extract.

[0055] The reaction mixture defined herein comprises an acid. Preferably, the acid is an inorganic acid or an organic acid, preferably an organic acid. Preferably, the organic acid is selected from acetic acid, ascorbic acid, lactic acid, malic acid, citric acid, succinic acid, tartaric acid, oxalic acid, tartronic acid, glycolic acid, glyceric acid, succinic acid, fumaric acid; sugar acids such as gluconic acid, glucuronic acid, glucaric acid, mannuronic acid; aromatic acids such as benzoic acid, p-hydroxybenzoic acid, catechin, gallic acid, ferulic acid, quinic acid; and mixtures thereof. Preferably, the organic acid is selected from ascorbic acid, lactic acid, malic acid and citric acid, succinic acid, tartaric acid, more preferably, the organic acid is selected from lactic acid, malic acid and citric acid, tartaric acid and combinations thereof, most preferably, the organic acid is lactic acid.

[0056] Preferably, the reaction mixture comprises at least 5 wt. % of acid based on the total weight of the reaction mixture, more preferably at least 10 wt. % based on the total weight of the reaction mixture.

[0057] In a preferred embodiment, step (i) comprises the step of adding an acid to the yeast extract. Preferably, the amount of acid added is 0.1-2 parts by weight of the acid added to 1-10 parts by weight of the yeast extract comprising at least 0.5% by weight of sugar amine and / or its derivatives based on the weight of the dry matter of the yeast extract.

[0058] The reaction mixture as defined herein comprises at least 10 wt% water, based on the weight of the reaction mixture. Preferably, the reaction mixture comprises at least 20 wt% water, preferably at least 30 wt% water, even more preferably at least 40 wt% water.

[0059] Typically, the reaction mixture is acidic so that a 1% by weight dilution in demineralized water has a pH of from 2.0 to 7.0, preferably from 3.0 to 6.5 and even more preferably from 4.0 to 5.5.

[0060] As defined herein, the method comprises at least 10 wt% water based on the weight of the reaction mixture. The water is provided by the yeast extract or is added separately. Preferably, the reaction mixture comprises at least 15 wt%, even more preferably at least 20 wt%, most preferably at least 25 wt% of the reaction mixture. The water may be provided by the yeast extract or is added separately.

[0061] In another preferred embodiment, the heated reaction mixture is acidic so that a 1 wt% dilution in demineralized water has a pH of 3.5 to 6.0, preferably 4.0 to 5.5, even more preferably 4.0 to 5.0. It has been found that when the pH is above 6, the off-flavors are stronger than the aromas.

[0062] In a preferred embodiment, the reaction mixture comprises:

[0063] a. 20-75% by weight of a yeast extract, which comprises at least 1% by weight of a sugar amine and / or a derivative thereof, based on the weight of the dry matter of the yeast extract, and at least 2% by weight of a thiol-containing compound, based on the weight of the dry matter of the yeast extract;

[0064] b. 2-20% by weight of the reaction mixture in an amount of an acid;

[0065] c. 10-80 wt % water based on the weight of the reaction mixture.

[0066] In some preferred embodiments, the method is carried out in the presence of a fat or oil. Where a fat is used, the fat is preferably a vegetable fat such as palm oil stearin, but fats from animals such as pigs, chickens and / or cattle are also within the scope of the invention. The fat preferably comprises triglycerides. Fat also means oils such as olive oil or palm oil.

[0067] In a preferred embodiment, the method according to the invention does not comprise the step of separately adding a reducing sugar. "Solely" means in addition to yeast extract. Reducing sugars are sugars containing a hemiacetal group or a hemiketal group. Examples of reducing sugars include fructose, glucose, ribose, xylose, maltodextrin, dextrin, glucose syrup.

[0068] Preferably, the reaction mixture is heated for a period of (t) hours according to the following formula (I):

[0069] T=4096e -bT (I)

[0070] Among them: 90℃≤T≤160℃, and 0.06≤b≤0.07.

[0071] The reaction mixture is preferably heated at a temperature of 90-160° C. for 0.05-20 hours, preferably 0.1-15 hours, more preferably 0.2-12 hours, even more preferably 0.5-10 hours, most preferably 1-8 hours.

[0072] In a preferred embodiment, the reaction mixture is heated at 90°C for at least 7.5 hours, preferably at least 11 hours, and preferably at most 18 hours.

[0073] In a preferred embodiment, the reaction mixture is heated at 100°C for at least 3.5 hours, preferably at least 6 hours, and more preferably at most 11 hours.

[0074] In a preferred embodiment, the reaction mixture is heated at 120°C for at least 0.75 hours, preferably at least 1.5 hours, and preferably at most 3.5 hours.

[0075] In a preferred embodiment, the reaction mixture is heated at 140°C for at least 0.15 hours, preferably at least 0.25 hours, and preferably at most 0.9 hours.

[0076] In a preferred embodiment, the reaction mixture is heated at 160°C for at least 0.04 hours, preferably at least 0.06 hours, and preferably at most 0.2 hours.

[0077] The reaction mixture is heated at a pressure sufficient to reach the desired temperature using methods known to the skilled person. In some embodiments, when the reaction mixture is heated at a temperature above 100° C., the reaction mixture is preferably heated in a container contacting an oil bath, in an autoclave, a pressure cooker, or in a continuous high temperature process. In some preferred embodiments, the reaction mixture is heated at a pressure of up to 4 bar. Typically, the reaction mixture is heated at 120° C. and 2 bar, and preferably at 140° C. and 3.6 bar.

[0078] Optionally, after the heating step, the reaction mixture obtained is subjected to a drying step c) by vacuum drying, spray drying or belt drying, optionally in the presence of a drying additive, preferably wherein the drying additive is a fat component, a starch component and / or a salt. Advantageously, the flavoring composition obtained after drying is easy to store, transport and use in food preparation. Typically, vacuum drying is carried out at a pressure as low as 10 mbar and a temperature of up to 80° C.

[0079] In a second aspect, the invention relates to a flavouring obtainable by a process as defined herein, wherein a 1% solution of the flavouring in demineralised water at 20°C has a pH of 3.5 to 6, preferably 4 to 5.

[0080] Preferably, the meaty aroma / flavor can be determined by sensory evaluation of an aqueous solution containing 0.01-0.5 wt. % by weight of dry matter of a flavoring obtainable by the process defined herein, preferably 0.02-0.2 wt. % by weight of dry matter of a flavoring obtainable by the process defined herein.

[0081] In a third aspect, the present invention relates to a flavouring composition comprising a yeast extract comprising, based on a dry yeast extract:

[0082] i. at least 20 mg / g of ribonucleotides,

[0083] ii. at least 20 mg / g of organic acid,

[0084] iii. at least 0.1 mg / g glutathione,

[0085] iv. preferably at least 1 mg / g of pyroglutamate,

[0086] v. preferably at least 1 mg / g of cyclo-cysteinyl-glycine,

[0087] vi. and wherein the composition is preferably free of thiamine and thiothiazol.

[0088] Preferably, a 1 wt% solution of the composition has a pH of 3.5 to 6 at 20°C.

[0089] In a fourth aspect, the present invention relates to a method for preparing a savoury food product comprising combining 0.001-1 % by weight of a flavouring composition prepared according to the method defined herein with one or more food ingredients.

[0090] In a fifth aspect, the present invention relates to the use of a composition to provide meat flavor to savory food, the composition comprising a yeast extract, the yeast extract comprising, based on a dry yeast extract:

[0091] i. at least 20 mg / g of ribonucleotides,

[0092] ii. at least 20 mg / g of organic acid,

[0093] iii. at least 0.1 mg / g glutathione,

[0094] iv. preferably at least 1 mg / g of pyroglutamate,

[0095] v. preferably at least 1 mg / g of cyclo-cysteinyl-glycine,

[0096] vi. and wherein the composition is preferably substantially free of thiamine and thiothiazol.

[0097] "Substantially free of thiamine" in the context of this application means less than 1 mg / g, preferably less than 0.1 mg / g, preferably less than 0.01 mg / g.

[0098] "Substantially free of thiothiazole" in the context of this application means less than 0.1 mg / g, and substantially free of thiothiazole, preferably less than 0.01 mg / g.

[0099] Preferably, the flavoring obtainable by the process defined herein comprises compounds such as 2-methyl-3-furanthiol, 2-methyl-3-(methylthio)furan and bis(2-methyl-3-furyl)disulfide.

[0100] As used herein, savory foods refer to foods such as soups, sauces, bouillons, soup cubes, meal packs, condiments and marinades.

[0101] The invention will now be illustrated by the following non-limiting examples.

[0102] Example

[0103] Materials and methods

[0104] Ingredients used:

[0105] Nuc-YEP: yeast extract powder, 12% ribonucleotides, 18% salt, purchased from Biospringer.

[0106] Yeast extract paste Maxarome from DSM, identified as Nuc-YE paste, was used.

[0107] GSH-YEP: yeast extract powder, 4% glutathione, <1% salt, purchased from Biospringer.

[0108] Lactic acid 80% (Corbion, NL),

[0109] HCl (Merck, DE)

[0110] Ascorbic acid (BASF, DE)

[0111] Malic acid (BASF, DE)

[0112] NMR quantitative measurements

[0113] qNMR analysis was performed according to SOP 890V1 (Quantitative NMR in Food Systems) using targeted profiling (Chenomx). 1D spectra were recorded on a Bruker Avance III 600 NMR spectrometer equipped with a 5 mm cryoprobe using the NOESYGPPR1D pulse sequence. 1 H NMR spectrum.The probe is tuned to detect the 1H resonance at 600.25MHz.The internal probe temperature is set to 298K.64 scans are collected at 57k data points, the relaxation delay is 10 seconds, the acquisition time is 4 seconds, and the mixing time is 100 milliseconds.Low power water suppression (16Hz) is applied for 0.99 seconds.Data are processed in TOPSPIN 3.5pl 1 version (BrukerBioSpin GmbH, Rheinstetten, Germany).The exponential window function is applied to free induction decay (FID) with a 0.15Hz line broadening factor before Fourier transform.Manual phase correction and baseline correction are applied to all spectra.The spectra are referenced for the methyl signal (δ0.0ppm) of TSP.

[0114] 1D 1 H NMR spectra were imported into Chenomx software (Chenomx NMR Professional Suite v8.13, Edmonton, Alberta, Canada). The relevant Chenomx model was fitted to the NMR signal of the target compound, minimizing the residual line. An internal (Matlab-based) reporting module calculated the compound concentration in the sample in three different units (i.e., % w / w, mg / g, and mg).

[0115] pH

[0116] A 1% solution of the reaction mixture was prepared in 5 g / L brine, and the pH was measured using a pH meter at 20°C.

[0117] SPME-GC / MS method

[0118] All GC-MS analyses were performed using an Agilent 6890GC coupled to a 5973MSD system equipped with a Gerstel multipurpose sampler (MPS2). Aromatic compounds were extracted from the samples by solid phase microextraction (SPME) using a polydimethylsiloxane absorbent fiber coating (PDMS). The column was a DB-WAX (20 m x 180 μm x 0.3 μm) from J&W Scientific.

[0119] Detailed conditions:

[0120] GC MS SPME method was used to analyze volatiles. The fiber was a PDMS 100 μm SPME fiber.

[0121] Oven: Start Temperature: 35°C (On) Maximum Temperature: 250°C; Start Time: 4.00 minutes Equilibration Time: 0.10 minutes Ramps: #Rate 1 Final Temperature / Final Time 5.00°C / minute 230°C 7.00 minutes Run Time: 50.00 minutes; Front Inlet (CIS3): Mode: Seamless; Start Temperature: 250°C (Off); Pressure: 132.4 kPa (On); Purge Flow: 50.0 mL / min; Purge Time: 1.00 minutes; Total Flow: 53.2 mL / min; Air Saver: Exhaust Type: Helium.

[0122] Column: capillary column; Model: J&W 121-7023DB-WAX;

[0123] Maximum temperature: 250℃ Nominal length: 20.0m; Nominal diameter: 180.00μm

[0124] Nominal film thickness: 0.30 μm; Mode: Constant flow rate; Initial flow rate: 1.0 mL / min

[0125] Nominal initial pressure: 132.5 kPa; Average velocity: 44 cm / sec; Inlet: front inlet

[0126] Inlet pressure: vacuum.

[0127] THERMAL AUX 2: Purpose: MSD transfer line heater; Description: MS-transfer; Start temperature: 250°C (on); GERSTEL CIS temperature: 250°C; GERSTEL MPS SPME injection.

[0128] Sample preparation: Incubation temperature: 60°C; Incubation time: 1.00 min; Stirrer speed: 500 rpm; Stirrer on time: 10 sec; Stirrer off time: 1 sec.

[0129] Sample parameters: Bottle penetration: 22.00 mm: Extraction time: 30.00 min

[0130] Injection penetration: 54.00 mm: Desorption time: 600 seconds; Cycle setting: Cycle time: 65.0 minutes.

[0131] MS acquisition parameters: solvent delay: 3.00 min; EM offset: 0

[0132] EM voltage: 1100; [Scan parameters] low quality: 29.0; high quality: 250.0

[0133] Threshold: 0; [MS area] MS four quadrants: 150℃ maximum 200℃.

[0134] Sensory

[0135] From a 1% solution of the reaction mixture in 5 g / L saline, dilutions of 0.2 g / L, 0.1 g / L, 0.05 g / L, and / or other dilutions as listed in the experiment were prepared using saline. The solutions were tasted by a group of three and evaluated for meaty flavor intensity and the presence of associated off-flavors. In Example 2, evaluations were also conducted in the presence of MSG (1 g / L). The results are recorded below:

[0136] +++ = strong meaty taste; ++ = meaty taste; + = weak meaty taste; - = no meaty taste

[0137] Example 1

[0138] 10 g Nuc-YEP, 1 g lactic acid (80%) and 10 g water were mixed and heated at 100°C for 1 hour. Subsequently, 1 g GSH-YEP was added, mixed and continued to heat at 100°C. 2 g (residue) was sampled at 1 hour, 2 hours, 3.5 hours, 5 hours and 7 hours. Store at -18°C. A 1% solution was made to measure pH, evaluate meaty taste, and further dilutions were made. Samples with a concentration of 0.1% by weight were prepared.

[0139] Table 1.1 Results of the reaction over time in Example 1

[0140] sample pH 0.100% by weight 1hr 4.95 - 7hr 4.8 +

[0141] Example 2

[0142] The same steps as in Example 1 were performed except that the mixture was heated at 100°C for 7 hours.

[0143] Table 2.1 Formula used

[0144] sample Nucl YEP 12% GSH-YEP-8% Lactic acid (80%) water 2.1 15 g 1.5g 1.5g 15g 2.2 15g 7.5g 1.5g 15g

[0145] Table 2.2 NMR analysis results of the mixture before and after heating

[0146]

[0147]

[0148] The sensory evaluation of the meaty taste and meaty aroma of the compositions obtained in Examples 2.1 and 2.2 was carried out on the following samples:

[0149] A = 0.1 wt. % composition in water.

[0150] B = 5 g sodium chloride / 0.1 wt% composition in L.

[0151] C = 0.1 wt% composition in 5 g sodium chloride / L + 1 g MSG / L.

[0152] Table 2.3 Sensory results of Example 2

[0153] sample A B C 2.1 + + + 2.2 ++ ++ ++

[0154] The measurements clearly show the relationship between the initial glutathione concentration and the intensity of the flavour obtained. The meaty flavour was detected in the presence of other food ingredients.

[0155] Example 3

[0156] The same procedure as Example 2 and Formulation 2.2 was used.

[0157] After 7 hours of reaction at 100°C, a sample was taken for tasting (Example 3.1). The remaining reaction mixture was dried at 60°C for 30 minutes and then at 80°C for 30 minutes in a vacuum oven. The powder was crushed in a mortar (3.2), and half of the powder was stored at -20°C and the other half at room temperature.

[0158] Table 3.1 Sensory results of Example 3

[0159] sample Taste, 0.1% in 5g salt / L 3.1(fuzzy) ++ 3.2(pink) +++

[0160] During drying, the meaty flavor is retained. Removal of moisture results in a powder that provides a more intense flavor (more flavor components per gram of product).

[0161] Example 4

[0162] The ratio of GSH-YEP:Nuc-YEP:lactic acid was varied at a constant concentration (50%). The reaction conditions were the same as in Example 2, ie, heating at 100°C for 7 hours.

[0163] Table 4.1 Recipes and results of reaction mixtures prepared with different component ratios

[0164]

[0165]

[0166] In the absence of lactic acid (4.11), no perceptible meaty flavor was detected.

[0167] Example 5

[0168] For Example 5, the optimum ratios in Examples 4.01 and 4.05 were used, while the amount of water was varied from 75% water addition to 200% water addition as shown in Table 5.1 below. The conditions were similar to those in Example 2, i.e. 7 hours at 100°C. The taste was corrected for the amount of water added in the experiment, corrected to 0.033 w / w based on the dry material concentration.

[0169] Table 5.1 Formulation and results of Example 5

[0170]

[0171] Yeast extract gave the best tasting results (5.04, 5.05, 5.06) at 40-50 wt% based on the total reaction mixture weight and a 2:1 ratio of GSH-YE to Nuc-YE.

[0172] Example 6

[0173] 6.1. 1 g Nuc-YEP, 1 g GSH-YX, 0.1 g lactic acid and 0.25 g water were mixed and heated at 120°C for 1 hour, allowed to cool, and a brown pasty solid with some sulfur smell was obtained. The 1% solution was slightly turbid, slightly yellow and smelled like cooked meat.

[0174] 6.2. As 6.1, but now with pre-reacted Nuc-YEP. 1 g Nuc-YEP, 0.1 g lactic acid and 0.25 g water were mixed and heated at 120°C for 15 seconds, then 1 g GSH-YE was added and heating continued at 120°C for 1 hour, allowed to cool. The mixture was a sticky, pasty brown mass with some sulfur smell.

[0175] 6.3. As 6.2, but now also 0.1 g of palm fat (saturated 49%, monounsaturated 37%, polyunsaturated 9%) flakes are added together with GSH_YEP.

[0176] sample pH Taste Results 6.1. 5.18 0.1 wt%++, 0.05 wt%++, 0.025 wt%+ 6.2. 5.42 0.1 wt%++, 0.05 wt%++, 0.025 wt%+ 6.3. 5.35 0.1 wt%++, 0.05 wt%++, 0.025 wt%+

[0177] Example 6.1 shows that reaction at 120° C. for 1 hour produces a tasting composition similar to that of reaction at 100° C. for 7 hours. Addition of palm fat resulted in meaty notes, but did not affect the organoleptic properties of the reaction mixture.

[0178] Reaction at 120°C for 1 hour was similar to reaction at 100°C for 7 hours. Nuc-YEP pre-reaction did not result in organoleptic differences, but different pH values ​​were obtained. Addition of palm fat resulted in meaty taste, but did not affect the organoleptic properties of the reaction mixture.

[0179] Example 7

[0180] Small amounts were mixed and heated in a Hungate tube tightly sealed with a butyl rubber septum in an oil bath at 120° C. for 1.5 hours, poured into a Petri dish to cool and evaporated to dryness in vacuo at 80° C. The residue was ground and crushed into a powder using a pestle and mortar (hot to avoid water absorption) and stored in a glass bottle under nitrogen.

[0181] 7.1.3g Biospringer GSH-YEP+1.5g Nuc-YEP+0.5g lactic acid 80%+5g water – mixture

[0182] 7.2.5g Biospringer GSH-YEP + 0.7g lactic acid 80% + 5g water – mixture

[0183] Table 7.1 pH and sensory results of the reaction mixture of Example 7

[0184] sample pH Taste Results 7.1. 4.64 0.05％+++ 7.2. 4.54 0.05％-

[0185] The lack of meaty flavor in sample 7.2 suggests that both sugar amines and sulfhydryl compounds are required to provide the desired flavor.

[0186] Example 8

[0187] Example 5 of JP2016-174587 was conducted as a comparative example. According to JP2016-174587, the following mixture was heated at 110°C for 30 minutes.

[0188] Table 8.1 Recipe and results

[0189]

[0190]

[0191] *FruGlu – Fructose / glucose obtained by mixing glucose and fructose 1:1 as a 50% solution in water.

[0192] Samples 8.1, 8.2 and 8.3 had a dominant umami taste. Samples 9.4 and 9.5 both had a meaty taste.

[0193] Example 9

[0194] Compositions heated at 100°C for 7 hours as in Example 5.06 with other acids or no acid (all powder dissolved within 1 hour) 30% GSH-YE (Biospringer), 15% Nuc-YE (Biospringer), 5% acid, 50% water. Tasting of 0.1% and 0.2% samples in 5g NaCl / L, n=3.

[0195] Table 9.1

[0196] sample acid pH, 1% in saline Sensory 0.1% All Sensory 0.2% 9.1 lactic acid 4.92 ++ +++ 9.2 ascorbic acid 5.60 ++ +++ 9.3 Malic Acid 4.57 ++ ++ 9.4 Hydrochloric acid (6M) 5.56 ++ ++

[0197] Example 10

[0198] Using a stainless steel tube tightly sealed with a screw-lined steel cap, the following mixture was heated at a specific temperature for a specific time, immersed in an oil bath.

[0199] Table 10.1

[0200]

[0201] Comparative Example A

[0202] Example 1 of JP 2007 259744A was carried out. 25 parts by weight of 5'-nucleotide yeast extract (product produced by Biospringer, yeast extract, 36% 5'-nucleotide), 10 parts by weight of glutathione-containing yeast extract (product produced by Kojin, yeast extract Aromild U-15, 15% glutathione), 0.2 parts by weight of ascorbic acid and 64.8 parts by weight of water were mixed. The mixture was heated at 105° C. for 40 minutes in a pressure reactor. The product was cooled and then tasted, and the results are shown in Table 11.

[0203] Comparative Example B

[0204] Example 1 of JP 2003 169627A was carried out. 7 parts by weight of a 5'-nucleotide yeast extract (product produced by Biospringer, yeast extract, 36% 5'-nucleotide content), 19 parts by weight of a glutathione-containing yeast extract (product produced by Kojin, yeast extract Aromild U-15, 15% glutathione content) and 65 parts by weight of water were mixed. The mixture was heated at 105° C. for 65 minutes in a sealed tube (pressurized), and then allowed to cool and then tasted (Table 11).

[0205] Comparative Example C

[0206] Example 5 of EP 2 103 225 A1 was carried out. The reaction contained nucleotides (but no yeast extract containing nucleotides), was reacted with acid (HCl) and heated. Then, yeast extract containing cysteine ​​was added, the pH was adjusted to 4.5 and heated at 95° C. for 5 hours. The total heating time was 22 hours.

[0207] Embodiment 5:

[0208] (1) 1.8 g IMP, 2.6 g H2O, 0.4 g concentrated HCl (4 mmol), 10 hours, 95°C

[0209] (2) Add 7.5g GSH-YE 4% BS + 0.6g glucose + 11.7g H2O

[0210] (3) Adjusting the pH to 4.5 with concentrated NaOH, and performing NMR determination on the drug to determine the % intact nucleotides (claim 1a)

[0211] (4) Heating for 5 hours at 95°C

[0212] (5) Adjust pH to 7.5 with concentrated NaOH

[0213] (6) Heating for 7 hours at 95°C

[0214] taste:

[0215] A solution of 0.2 g of A, B and C and 0.5 g of salt was made in warm water and tasted. 3 persons (n=3) compared this with 0.2 g of the mixture prepared according to the invention (as in Example 7.1) and 0.5 g of salt in 100 mL of warm water.

[0216]

[0217]

[0218] *1 wt. % dilution in demineralized water at 20°C.

Claims

1. A method for preparing a flavoring agent, comprising the following steps: (i) providing a reaction mixture, the reaction mixture comprising: a. a yeast extract, the yeast extract comprising at least 0.5% by weight of sugar amines and / or derivatives thereof, based on the weight of the dry matter of the yeast extract, and at least 1.0% by weight of thiol-containing compounds, based on the weight of the dry yeast extract; b. at least 2% by weight of an inorganic or organic acid based on the weight of the reaction mixture; c. at least 10% by weight of water, based on the weight of the reaction mixture; (ii) heating the reaction mixture at a temperature of 90-160° C., wherein the heated reaction mixture is acidic so that a 1 wt % dilution in demineralized water has a pH of 3.5 to 6 at 20° C.; (iii) optionally drying the product, wherein the reaction mixture is heated for a period of (t) hours according to the following formula: t=4096e -bT ,in: 90°C≤T≤160°C, and 0.06≤b≤0.07 (Formula I).

2. The method according to claim 1, wherein step (i) comprises providing a first yeast extract and a second yeast extract, wherein the first yeast extract comprises at least 0.5 wt. % of sugar amines and / or derivatives thereof, based on the weight of dry matter of the first yeast extract, and the second yeast extract comprises 1 wt. % of sulfhydryl-containing compounds, based on the weight of dry matter of the second yeast extract.

3. The method of claim 1, wherein step (i) comprises providing a yeast extract comprising at least 1.0 wt. % of sugar amine and / or its derivatives based on the weight of dry matter of the yeast extract and at least 1 wt. % of sulfhydryl-containing compounds based on the weight of dry matter of the yeast extract.

4. The method according to any one of the preceding claims, wherein the sugar amine and / or its derivative contains a ribose moiety, preferably wherein the sugar amine and / or its derivative is a ribonucleotide and / or a ribonucleoside.

5. The method according to any one of the preceding claims, wherein the thiol-containing compound is glutathione or cysteine, preferably glutathione.

6. The process according to any one of the preceding claims, wherein the reaction mixture comprises at least 2% by weight of an organic acid, preferably an organic acid selected from lactic acid, malic acid and citric acid, tartaric acid.

7. The process according to any one of the preceding claims, wherein the reaction mixture comprises at least 5 wt.-% of an inorganic or organic acid, based on the total weight of the reaction mixture.

8. The process according to any one of the preceding claims, wherein the reaction mixture comprises at least 20 wt. % water, preferably at least 30 wt. % water, even more preferably at least 40 wt. % water, based on the weight of the reaction mixture.

9. A process according to any one of the preceding claims, wherein the reaction mixture in step (ii) is acidic such that a 1 wt% dilution in demineralised water has a pH of 4 to 5.

10. The process according to any one of the preceding claims, wherein the reaction mixture is heated at a temperature of 90-160°C for a period of 0.05-20 hours, preferably 0.1-15 hours, more preferably 0.2-12 hours, even more preferably 0.5-10 hours, most preferably 1-8 hours.

11. The process according to any one of the preceding claims, wherein step c) is performed by vacuum drying, spray drying or belt drying, optionally in the presence of a drying additive, preferably wherein the drying additive is a fat component, a starch component or a salt.

12. Natural flavoring obtainable by the process according to any one or more of claims 1 to 11.

13. The natural flavoring composition according to claim 12, comprising a yeast extract, wherein the yeast extract comprises: i. at least 20 mg / g of ribonucleotides, ii. at least 20 mg / g of organic acid, iii. at least 0.1 mg / g glutathione (reduced and oxidized combined), iv. preferably at least 1 mg / g of pyroglutamate, v. preferably at least 1 mg / g of cyclo-cysteinyl-glycine, And wherein a 1 wt% solution of the composition has a pH of 3.5 to 6 at 20°C.

14. The natural flavoring composition according to claim 12 or 13, wherein the composition is substantially free of thiamine and thiothiazole.

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