WO2015021823A1 - 脂肽类化合物用于改善饮料酒香气的用途及其测定方法 - Google Patents
脂肽类化合物用于改善饮料酒香气的用途及其测定方法 Download PDFInfo
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- WO2015021823A1 WO2015021823A1 PCT/CN2014/080702 CN2014080702W WO2015021823A1 WO 2015021823 A1 WO2015021823 A1 WO 2015021823A1 CN 2014080702 W CN2014080702 W CN 2014080702W WO 2015021823 A1 WO2015021823 A1 WO 2015021823A1
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- compound
- liquor
- lipopeptide
- lichenin
- aroma
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Classifications
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L2/00—Non-alcoholic beverages; Dry compositions or concentrates therefor; Their preparation
- A23L2/52—Adding ingredients
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L2/00—Non-alcoholic beverages; Dry compositions or concentrates therefor; Their preparation
- A23L2/52—Adding ingredients
- A23L2/56—Flavouring or bittering agents
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L27/00—Spices; Flavouring agents or condiments; Artificial sweetening agents; Table salts; Dietetic salt substitutes; Preparation or treatment thereof
- A23L27/88—Taste or flavour enhancing agents
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12G—WINE; PREPARATION THEREOF; ALCOHOLIC BEVERAGES; PREPARATION OF ALCOHOLIC BEVERAGES NOT PROVIDED FOR IN SUBCLASSES C12C OR C12H
- C12G3/00—Preparation of other alcoholic beverages
- C12G3/04—Preparation of other alcoholic beverages by mixing, e.g. for preparation of liqueurs
- C12G3/06—Preparation of other alcoholic beverages by mixing, e.g. for preparation of liqueurs with flavouring ingredients
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/88—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86
- G01N2030/8809—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86 analysis specially adapted for the sample
- G01N2030/8813—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86 analysis specially adapted for the sample biological materials
Definitions
- lipopeptide compounds for improving aroma of beverage wines and method for determining the same
- the invention belongs to the field of food, relates to the use of a lipopeptide compound for improving the aroma of a beverage, a preparation method thereof, a method for the determination thereof, and a production process of a beverage wine. Background technique
- Matrix compounds in foods affect the aroma and quality of foods.
- the migration of aroma from food to the consumer's olfactory organ is a process that is influenced by many factors, including chemical interactions between food and aroma components, physical factors, consumer physiological factors (chewing time, efficiency, breathing).
- olfactory receptors Considering that sensory perception is an important factor in determining consumer preferences, understanding how aroma substances are released from food and interacting with olfactory receptors is a very important topic in food chemistry research.
- Classical studies abroad such as ⁇ -lactoglobulin in fresh milk, ⁇ -cyclodextrin in colloids, and phenolic compounds in wine. At present, there is little research on matrix compounds in white spirits, especially Chinese liquors.
- Ultra-high performance liquid chromatography coupled with mass spectrometry is an important means of studying macromolecular matrix compounds.
- Headspace solid phase microextraction HS-SPME
- GC-MS gas chromatography-mass spectrometry
- Lipopeptides are an important class of compounds produced by Bacillus, including Surfactin, Lichenysin, Itiirin, Fengycin, etc., which have antibacterial, anticancer and The important effect of anti-inflammatory.
- Surface is reported to be a powerful surfactant that reduces the surface tension of water from 70 niN/ni to 36 mN/m and a critical micelle concentration of 15.6 m g /L. Iturin is now reported to be a strong antibiotic with strong inhibitory properties against yeast and mold in fungi. Its inhibitory properties and surface properties show enhanced inhibition properties.
- Fengycin now reported to be an antibiotic mold suppression, mainly for plants to inhibit pathogenic bacteria fog 0
- One aspect of the invention relates to a method of improving or enhancing the aroma of a beverage comprising the step of using one or more effective amounts of a lipopeptide compound.
- the amount of the lipopeptide compound is greater than lg/L, greater than 10 g/L, greater than 100 ⁇ g/L, 1 to 1000 g/L, 1 to 500 g/L, 1 to 200 g/L, and 1-100.
- g/L 1 to 10 g/L, 10 to 1000 g/L, 10 to 500 g/L, 10 to 200 g/L, 10 to 100 g/L, 100 to 1000 g/L, 100 to 900 g /L, 100-500 g/L, 100-200 g/L or 200-500 g/L.
- beverages Choinese liquor, rice wine, wine, beer, sake, shochu, whiskey, brandy, vodka, rum, gin
- the total content of lignin after the addition is 500-1500 g/L, preferably 600-1400 g/L, 700-1300 g/L, 800-1200 g/L, 900-1100 g/L, in particular It is preferably 950 to 1050 g/L, for example, 1000 g/L.
- the content of lichenin in the drinking wine can be determined by the method of the present invention.
- Another aspect of the invention relates to a process for the preparation of a lipopeptide compound, characterized in that it is isolated and purified using white spirit;
- the beverage wine purified by the SPE C18 cartridge is purified by using a preparative liquid phase
- a further aspect of the invention relates to a method for determining the content of lipopeptides, particularly lichenin, in beverages
- the method is a combination of ultra performance liquid chromatography and mass spectrometry.
- Ultra-high performance liquid chromatography parameters UPLC TM BEH C 18 column (100 mm x 2.1 mm id, 1.7 ⁇ ), column temperature 30, injection volume 2 ⁇ , flow rate 0.3 mL/min, mobile phase A: 0.1 % methanolic acid solution, B: methanol; elution procedure: 0—0.1 min, 15% B; 0.1-10 min, 0% B; 10-11 min, 15% B;
- a further aspect of the invention relates to a composition comprising one or more lipopeptide compounds; optionally, the composition further comprises a pharmaceutically or food acceptable carrier or adjuvant;
- the composition is a food product, in particular a drink.
- the content of the lipopeptide compound is greater than tg/L, greater than 10 g/L, greater than 100 g/L, 1 to 1000 g/L, 1 to 500 g/L, 1 to 200 g/L, 1— 100 g/L, 1-10 g/L, 10-1000 g/L, 10-500 g/L, 10-200 g/L, 10-100 g/L, 100-1000 g/L, 100-900 g/L, 100-500 g/L, 100-200 g/L or 200-500 g/L.
- the composition is a beverage, consider the possible lignin content of the beverage itself.
- the total content of the lignin after the addition is 500-1500 g/L, preferably 600-1400 g/L, 700-1300 g/L, 800-1200 g/L, 900-1100 g/L, in particular It is preferably 950 to 1050 g/L, for example, 1000 g/L.
- the content of lichenin in the beverage wine can be determined by the method of the present invention.
- a further aspect of the invention relates to the use of a lipopeptide compound for the preparation or production of a beverage.
- the amount of the lipopeptide compound is greater than lg/L, greater than 10 g/L, greater than 100 g/L, 1 to 1000 g/L, 1 to 500 g/L, 1 to 200 g/L, 1— 100 g/L, l-10 g/L, 10-1000 g/L, 10-500 g/L, 10-200 g/L, 10-100 g/L, 100-1000 g/L, 100-900 g/L, 100-500 g/L, 100-200 g/L or 200-500 g/L.
- the drink is white wine
- the possible lignin content of the liquor itself should be considered.
- the total content of lignin after the addition is 500-1500 g/L, preferably 600-1400 g/L, 700-1300 g/L, 800-1200 g/L, 900-1100 g/L, in particular It is preferably 950 to 1050 g/L, for example, 1000 g/L.
- the content of lichenin in the liquor can be determined by the method of the present invention.
- a further aspect of the invention relates to the use of a lipopeptide compound for improving or enhancing the aroma of a beverage.
- the amount of the lipopeptide compound is greater than lg/L, greater than 10 g/L, greater than 100 g/L, 1 to 1000 g /L, 1 to 500 g/L, 1 to 200 g/L, 1— 100 g / L, 1 - 10 g / L, 10 - 1000 g / L, 10 - 500 g / L, 10 - 200 g / L, 10 - 100 g / L, 100 - 1000 g / L, 100 - 900 g/L, 100-500 g/L, 100-200 g/L or 200-500 g/L.
- a further aspect of the invention relates to the use of a lipopeptide compound for increasing the volatility of aroma compounds and/or reducing the volatility of a odorant compound in a beverage; in particular, the aroma compounds are in esters, alcohols and acid compounds Any one or more of the odorant compounds is any one or more of phenol compounds.
- the amount of the lipopeptide compound is greater than lg/L, greater than 10 g/L, greater than 100 g/L, 1 to 1000 g /L, 1 to 500 g/L, 1 to 200 g/L, 1— 100 g / L, 1 - 10 g / L, 10 - 1000 g / L, 10 - 500 g / L, 10 - 200 g / L, 10 - 100 g / L, 100 - 1000 g / L, 100 - 900 g/L, 100-500 g/L, 100-200 g/L or 200-500 g/L.
- the effect of the lipopeptide compound on the volatile aroma component is determined by headspace solid phase microextraction-gas chromatography (HS-SPME-GC-MS); specifically, the steps are as follows:
- headspace solid phase microextraction 8 mL of the sample was added to a 20 mL headspace vial for headspace solid phase microextraction.
- the conditions of headspace solid phase microextraction are: three-phase extraction head (DV B/CAR/PDMS, 50/30 ⁇ ), preheating 5 niin at 20-30 °C, extraction adsorption 30-50 niin, GC desorption for 5 min.
- GC condition column DB-Wax (60 mxO.325 mmx0.25 ⁇ ); Temperature program: 2 min at 50 °C, 230 °C at 6 °C/min for 15 min; inlet temperature 250 V, carrier gas He, flow rate 2 mL/min; no splitting.
- a further aspect of the invention relates to a beverage wine production process comprising the step of using one or more lipopeptide compounds.
- the amount of the lipopeptide compound is greater than lg/L, greater than 10 g/L, greater than 100 g/L, 1 to 1000 g/L, 1 to 500 g/L, 1 to 200 g/L, 1— 100 g/L, 1-10 g/L, 10-1000 g/L, 10-500 g/L, 10-200 g/L, 10-100 g/L, 100-1000 g/L, 100-900 g/L, 100-500 g/L, 100-200 g/L or 200-500 g/L.
- the possible lichenilin content of the beverage When added to a beverage, consider the possible lichenilin content of the beverage itself.
- the total content of lignin after the addition is 500-1500 g/L, preferably 600-1400 g/L, 700-1300 g/L, 800-1200 jig/L, 900-1100 jig/L, in particular
- the content of melamine in 950-1050 jig/L, for example 1000 jig/Lo beverage can be determined by the method of the present invention.
- the present invention can be determined by the method of the present invention.
- the lipopeptide compound is selected from any one or more selected from the group consisting of Surfactin, Lichenysin, Iturins, and Fengycin, unless otherwise specified.
- the lichenin is any one or more of the following homologues selected from the group consisting of the following formulae (I)-(VII):
- the mass-to-charge ratio (ni/z) of the above formulas (I) to (VII) is 993, 1007, 1021, 1035, 1049, 1007, and 1021, respectively.
- the beverage wine flavoring compound includes an aroma compound and a hetero-olfactory compound, and if not specified, the aroma compound refers to any one or more of an ester, an alcohol, and an acid compound, and the hetero-olfactory compound refers to Any one or more of phenolic compounds.
- the esters, alcohols, and acid compounds or phenolic compounds are esters, alcohols, and acid compounds or phenolic compounds in beverages, particularly Chinese beverages.
- the food is preferably a drink; specifically, the drink is alcohol (Chinese liquor, rice wine, wine, beer, sake, shochu, whiskey, brandy, vodka, rum, gin). More specifically, the wine is white wine such as Chinese white wine; further specifically, the Chinese white wine is a sauce-flavored, rich-flavored, fragrant, musk-type or medicinal liquor.
- the present inventors used HS-SPME to study the volatile effects of lipopeptide compounds (lichenin) on aroma compounds in beverage wines. The extraction temperature and extraction time were determined at 20-30 Torr and 30-50 min. Lipopeptides, especially lichenin, have a significant functional effect on the volatility of aroma compounds in various beverages. Lipopeptides, especially lichenin, have a significant effect on esters, alcohols, acids and phenols in wine.
- Figure 1 Effect of extraction time on the volatility of aroma compounds (esters, alcohols and acids) and iso- olfactory compounds (phenols) in white spirits.
- Figure 2 Elution profile of the separation of lichenin. (Each peak corresponds to a homolog).
- Figure 3 MS/MS spectrum of lichenin (mass-to-charge ratio 1035.6).
- Figure 4 1H nuclear magnetic spectrum (Figure 4A) and 13 C nuclear magnetic spectrum (Figure 4B) of her coat (molecular weight 1034.6). detailed description
- Lichenin (Formula I-Formula VI I Mixture): It may be commercially available or prepared according to the records in the literature disclosed in the prior art, or may be prepared by the method of Example 4 below. This example specifically uses the lichenin prepared according to Example 4, which is a mixture of seven homologues (seven homologs are mixed in equal proportions).
- Lichenin Commercially available or referenced to prior art such as: Yakimov, MM; Abraham, W.-R.; Meyer, H.; Giuliano, L.; Golyshin, PN, Structural characterization of lichenysin A components by Fast atom bombardment tandem mass spectrometry. Biochimica et Biophysica Acta 1999, 1438 (2), 273-280.
- Lichenin Commercially available or referenced to prior art such as: Yakimov, MM; Abraham, W.-R.; Meyer, H.; Giuliano, L.; Golyshin, PN, Structural characterization of lichenysin A components by Fast atom bombardment tandem mass spectrometry. Biochimica et Biophysica Acta 1999, 1438 (2), 273-280.
- Liquor liquor sample Langjiu wine sample (Before adding, the content of lichenin in the wine sample is 100 g / L ; added to 1000
- the sensory evaluation method is as follows:
- the wine sample was judged by 20 national liquor judges at room temperature.
- the wine sample is diluted to the same degree (for example, 55 degrees, less than 55 degrees can be diluted) for evaluation.
- the sensory evaluation scored 110 points.
- the scores were evaluated in five aspects: color, fragrance, taste, style and empty cup.
- the difference in wine quality was 10 points, 25 points, 50 points, 15 points. 10 points; For the empty cup incense, put it for 30 minutes after being dried and compare the intensity.
- Example 2 Optimization of SPME extraction conditions in a simulated system
- HS-SPME-GC-MS was used to study the volatility of several aroma compounds (esters, alcohols and acids) and iso- odor compounds (phenols) in the simulated system. Firstly, the experimental and conditional parameters of HS-SPME were determined. , as follows-
- Headspace solid phase microextraction The conditions are as follows: Three-phase extraction head (DV B/CAR/PDMS, 50/30 ⁇ ), preheated at 20-30 °C for 5 min, extracted for 30-50 min, and desorbed by GC for 5 min.
- GC condition column DB-Wax (60 mxO.325 mmx0.25 ⁇ ); temperature program: hold at 50 °C for 2 min, increase to 230 °C at 6 °C/min for 15 min; inlet Temperature 250 °C, carrier gas He, flow rate 2 niL/miii; no splitting.
- Liquor liquor sample 1 1-3 They are liqueur liquor, fragrant wine, and fragrant wine.
- lichenin has a significant functional effect on the volatility of aroma compounds such as esters, alcohols, acids, and odorous compounds in alcohol.
- Lichenin inhibits the volatilization of phenols and promotes the volatilization of esters, alcohols and acids.
- the volatilization of the olfactory compound phenolic compound is significantly reduced, that is, lichenin is beneficial to reduce the volatilization amount of the olfactory compound in the liquor, and the reduction is between 50% and 70%.
- Example 4 Effect of lichens 3 ⁇ 4 on the volatility of aroma compounds in wine and beer
- the lipopeptides in the liquor were purified using SPE C18 cartridges (Waters) and the extraction equipment was waters 2767 Sample Manager, Waters 2489 UV/visible Detector, and Waters 2535 Quaternary Gradient Module.
- Mass spectrometry of lichenin in liquor was performed using aters Acquity UPLC and Synapt Q-TOF system.
- Sauce-flavored liquor (Langjiu) The sample purified by SPE C18 cartridge was purified by preparative liquid phase. The column was analyzed on a Xbridge Prep C 18 (250 mm x 19 mm, 10 ⁇ ) reversed-phase column at 210 nm. The mobile phase ratio for separation is: 0.1% formic acid in methanol (A), methanol (B). One separation procedure: 0-10 min, 40% B - 25% B; 10 - 50 min, 25% B - 5% B; 50 - 55 min, 5% B - 0% B. The sample was concentrated by evaporation to give a pure product.
- Mass Spectrometry Detector (MSD) Parameters Positive ion mode, atomization pressure 30 psi; separation voltage 3.5 KV, induced collision voltage 20,, dry gas N 2 , drying temperature 100 V, ion scanning range 50-1500 m/z. The data was collected and analyzed using the MaSS lyn X 4.1 software.
- Lichenin was isolated and purified from liquor. Surface elements, iturin and amylin can also be isolated. Among them, the lichenin product obtained seven homologous compounds, respectively. As can be seen from Figure 2, after preparation, 993.6 (671.4), 1007.6 (685.4, 671.4), 1021.6 (685.4, 671.4), 1035.6 (685.4), 1049.6 were obtained.
- Mass spectrometry analysis of compounds with a mass-to-charge ratio of 1035.6 (685.4) was carried out by Q-TOF-MS.
- the amino acid segment of the lichenin is basically consistent with the lichens reported in the literature.
- the amino acid segment is N,-Gln-Leu-Leu-Val-Asp-Leii-Ile-C', and the fatty acid segment is C15. fatty acid.
- the 1 H and 13 C NMR spectra were analyzed by a nuclear magnetic resonance spectrometer (Bruker AM 600).
- the solvent was deuterated methanol and the experimental temperature was 323 K.
- a chemical shift of CH—NH— which is also an important functional group that forms a cyclic peptide.
- the cyclolipopeptide must be formed from 7 amino acids and 1 hydroxy fatty acid.
- the information in the 13 C spectrum of Table 5 basically reflects the structural information of the compound.
- the volatile macromolecular compound in the wine is a lipopeptide compound, including surface element, lichenin, iturin and amylin.
- the content of the wine was calculated from the standard curve in Table 6.
- Table 8 Lichenin content in several major flavor liquors (g/L) (LC-MS/MS)
- the detection limit is basically above 1 g/L, which greatly improves the detection sensitivity of lichenin.
- Existing diode array detection methods generally have a detection concentration of 10 mg/L or more.
- the content of lichenin in liquor is between 10 and 100 g/L, but the content of lichenin varies greatly between samples.
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Abstract
脂肽类化合物用于改善或提高饮品香气的用途,所述饮品为酒类,例如中国白酒。制备脂肽类化合物的方法,其特征在于使用白酒进行分离纯化。测定白酒中脂肽类化合物含量的方法,其为超高效液相色谱与质谱联用方法。所述脂肽类化合物选自表面素、地衣素、伊枯草菌素和丰原素。
Description
脂肽类化合物用于改善饮料酒香气的用途及其测定方法 技术领域
本发明属于食品领域, 涉及脂肽类化合物用于改善饮料酒香气的用途、 其制备方 法、 其测定方法, 以及一种饮料酒生产工艺。 背景技术
食品中的基质化合物, 影响着食品的香气感官与品质。 香气从食品中迁移至消费 者的嗅觉器官是一个受众多因素影响的过程, 包括食品与香气成分之间的化学相互作 用, 物理学因素, 消费者生理因素 (咀嚼时间, 效率, 呼吸)等。 考虑到感官感受是决定 消费者喜好的重要因素, 因此了解香气物质如何从食品释放并与嗅觉受体作用是食品 化学研究中一个非常重要的课题。 国外的经典研究, 如鲜奶中的 β-乳球蛋白, 胶体中 的 β-环糊精, 葡萄酒中的酚类化合物。 目前对白酒特别是中国白酒中基质化合物的研 究甚少。
现如今, 饮料酒的品质主要关注于香气化合物的鉴定及定量, 但光是香气化合物 不能反映酒的香气感官, 其中基质化合物决定影响着酒的香气品质。 超高效液相色谱 联用质谱技术 (UPLC— MS), 是现在研究大分子基质化合物的重要手段。 顶空固相微 萃取 (HS— SPME)和气相色谱质谱 (GC— MS)是现在研究大分子基质化合物对香气影 响的重要手段。 饮料酒中挥发性成分的研究较多, 但对其非挥发性成分-大分子化合物 的研究相对较少。
脂肽类化合物是芽孢杆菌产生的一类重要化合物, 包括表面素 (Surfactin) 、 地 衣素 ( Lichenysin ) 、 伊枯草菌素 ( Itiirin ) 、 丰原素 ( Fengycin ) 等, 其具有抗菌、 抗癌和消炎的重要功效。 表面素现报道是一种强力的表面活性剂, 可以将水的表面张力 从 70 niN/ni降低至 36 mN/m, 临界胶束浓度为 15.6 mg/L。 伊枯草菌素现报道是一种较 强的抗生素, 对真菌中的酵母和霉菌具有较强的抑制性能。 其抑制性能, 与表面素表现 出增强抑制性能。 丰原素现报道是一种抑制霉菌的抗生素, 主要用于植物中的抑制致病 雾菌 0
然而, 目前尚没有关于脂肽类化合物 (例如地衣素) 改善饮品特别是饮料酒的香气
的相关报道。 发明内容
本发明人经过深入的研究和创造性的劳动, 发现脂肽类化合物例如表面素 (Surfactin) 、 地衣素 ( Lichenysin ) 、 伊枯草菌素 (Iturins) 和丰原素 (Fengycin) 等, 对于提升饮料酒的品质特别是改善香气发挥了重要作用。 由此提供了下述发明: 本发明的一个方面涉及一种改善或提髙饮品香气的方法, 包括使用一种或者多种 有效量的脂肽类化合物的步骤。 具体地, 脂肽类化合物的用量为大于 l g/L 、 大于 10 g/L 、 大于 lOOjig/L 、 1— 1000 g/L、 1— 500 g/L、 1— 200 g/L、 1— 100 g/L、 1一 10 g/L、 10— 1000 g/L、 10— 500 g/L、 10— 200 g/L、 10— 100 g/L、 100— 1000 g/L、 100-900 g/L、 100-500 g/L、 100— 200 g/L或者 200— 500 g/L。 当添加到饮料酒 (中国白酒、 黄酒、 葡萄酒、 啤酒、 清酒、 烧酒、 威士忌、 白兰 地、 伏特加、 朗姆、 金酒) 中时, 要考虑饮料酒自身中可能的地衣素含量。 优选地, 添加后的地衣素总的含量为 500— 1500 g/L,优选为 600— 1400 g/L、700— 1300 g/L、 800— 1200 g/L、 900— 1100 g/L, 特别优选为 950— 1050 g/L , 例如 1000 g/L。 饮 料酒中地衣素的含量可以采用本发明中的方法测定。 本发明的另一方面涉及一种制备脂肽类化合物的方法, 其特征在于使用白酒进行 分离纯化;
具体地,
将经过 SPE C18小柱净化后的饮料酒, 釆用制备液相进行纯化;
更具体地,
色谱柱釆用 Xbridge Prep C18 (250 mm x 19 mm, 10 μιη) 反相色谱柱, 210 nm 检测; 分离采用的流动相配比为: 0.1%的甲酸甲醇溶液 (A) , 甲醇(B) ; 一次分离 程序: 0— lOmin, 40 B-25%B; 10— 50 min, 25%B-5%B; 50— 55 min, 5%B— 0%B。 本发明的再一方面涉及一种测定饮料酒中脂肽类化合物特别是地衣素含量的方
法, 其为超高效液相色谱与质谱联用方法。
具体地,
超高效液相色谱参数: 色谱柱采用 UPLCTMBEH C18 柱 (100 mm x 2.1 mm i.d., 1.7 μηι ) , 柱温 30 , 进样体积 2 μί, 流速为 0.3 mL/min, 流动相 A: 0.1%的甲酸 甲醇溶液, B: 甲醇; 洗脱程序: 0— 0.1 min, 15%B; 0.1— 10 min, 0%B; 10— 11 min, 15%B;
质谱检测器 (MSD) 参数: 正离子模式, 雾化气压 30psi; 分离电压 3.5KV, 诱 导碰撞电压 20 V, 干燥气体 N2, 干燥温度 100°C, 离子扫描范围 50— 1500 m/z。
应用 Masslynx 4.1 软件对数据进行采集和分析。 本发明的再一方面涉及一种组合物, 其含有一种或多种脂肽类化合物; 可选地, 所述组合物还含有药学上或者食品学上可接受的载体或者辅料; 具体地, 所述组合物 为食品特别是饮品。 具体地, 脂肽类化合物的含量为大于 tg/L 、 大于 lO g/L 、 大 于 100 g/L 、 1— 1000 g/L、 1— 500 g/L、 1— 200 g/L、 1— 100 g/L、 1— 10 g/L、 10-1000 g/L、 10-500 g/L、 10— 200 g/L、 10— 100 g/L、 100— 1000 g/L、 100 -900 g/L、 100-500 g/L、 100— 200 g/L或者 200— 500 g/L。 当组合物为饮料酒时, 要考虑饮料酒自身中可能的地衣素含量。 优选地, 添加后 的地衣素总的含量为 500— 1500 g/L, 优选为 600— 1400 g/L、 700— 1300 g/L、 800 — 1200 g/L、 900— 1100 g/L, 特别优选为 950— 1050 g/L , 例如 1000 g/L。 饮料 酒中地衣素的含量可以采用本发明中的方法测定。 本发明的再一方面涉及脂肽类化合物在制备或生产饮品中的用途。 具体地, 脂肽 类化合物的用量为大于 l g/L 、大于 lO g/L 、大于 lOO g/L 、 1— 1000 g/L、 1— 500 g/L、 1— 200 g/L、 1— 100 g/L、 l— 10 g/L、 10— 1000 g/L、 10— 500 g/L、 10— 200 g/L、 10— 100 g/L、 100— 1000 g/L、 100— 900 g/L、 100— 500 g/L、 100— 200 g/L或者 200— 500 g/L。 本发明中, 当饮品为白酒时, 要考虑白酒自身中可能的地衣素含量。 优选地, 添 加后的地衣素总的含量为 500— 1500 g/L, 优选为 600— 1400 g/L、 700— 1300 g/L、 800— 1200 g/L、 900— 1100 g/L, 特别优选为 950— 1050 g/L , 例如 1000 g/L。 白 酒中地衣素的含量可以采用本发明中的方法测定。
本发明的再一方面涉及脂肽类化合物在改善或提高饮品香气中的用途。 具体地, 脂肽类化合物的用量为大于 l g/L 、 大于 lO g/L 、 大于 lOO g/L 、 1— 1000 g/L、 1 — 500 g/L、 1— 200 g/L、 1— 100 g/L、 1— 10 g/L、 10— 1000 g/L、 10— 500 g/L、 10— 200 g/L、 10— 100 g/L、 100— 1000 g/L、 100— 900 g/L、 100— 500 g/L、 100 -200 g/L或者 200— 500 g/L。
本发明的再一方面涉及脂肽类化合物在提高饮品中香气化合物挥发性和 /或降低 异嗅化合物挥发性的用途; 具体地, 所述香气化合物为酯类、 醇类和酸类化合物中的 任意一种或者多种, 所述异嗅化合物为苯酚类化合物中的任意一种或者多种。具体地, 脂肽类化合物的用量为大于 l g/L 、 大于 lO g/L 、 大于 lOO g/L 、 1— 1000 g/L、 1 — 500 g/L、 1— 200 g/L、 1— 100 g/L、 1— 10 g/L、 10— 1000 g/L、 10— 500 g/L、 10— 200 g/L、 10— 100 g/L、 100— 1000 g/L、 100— 900 g/L、 100— 500 g/L、 100 -200 g/L或者 200— 500 g/L。
在本发明的一个实施方案中, 采用顶空固相微萃取一气质连用(HS— SPME— GC 一 MS) 法测定脂肽类化合物对挥发性香气成分的影响; 具体地, 步骤如下:
20 mL顶空瓶中加入 8 mL样品, 进行顶空固相微萃取。 顶空固相微萃取的条件 为: 三相萃取头 ( DV B/CAR/PDMS, 50/30 μηι ) , 20— 30 °C预热 5 niin, 萃取吸附 30 -50 niin, GC解吸 5min。 GC条件 色谱柱: DB-Wax (60 mxO.325 mmx0.25 μηι ); 升温程序: 50 °C保持 2min, 以 6 °C/min的速率升至 230 °C, 保持 15 min; 进样口温 度 250 V,载气 He,流速 2 mL/min;不分流。 MS条件 EI电离源,离子源温度 230 °C, 电子能量 70eV, 扫描范围 35.00— 350.00 amu。 本发明的再一方面涉及一种饮料酒生产工艺, 包括使用一种或多种脂肽类化合物 的步骤。具体地, 脂肽类化合物的用量为大于 l g/L 、大于 lO g/L 、大于 lOO g/L 、 1— 1000 g/L、 1— 500 g/L、 1— 200 g/L、 1— 100 g/L、 1— 10 g/L、 10— 1000 g/L、 10-500 g/L、 10-200 g/L、 10— 100 g/L、 100— 1000 g/L、 100— 900 g/L、 100 -500 g/L、 100-200 g/L或者 200— 500 g/L。 当添加到饮料酒中时, 要考虑饮料酒自身中可能的地衣素含量。 优选地, 添加后 的地衣素总的含量为 500— 1500 g/L, 优选为 600— 1400 g/L、 700— 1300 g/L、 800 -1200 jig/L, 900-1100 jig/L, 特别优选为 950—1050 jig/L , 例如 1000 jig/Lo 饮料 酒中她衣素的含量可以采用本发明中的方法测定。
本发明中,
所述脂肽类化合物, 如果没有特别说明, 选自表面素 ( Surfactin ) 、 地衣素 (Lichenysin)、伊枯草菌素(Iturins)和丰原素(Fengycin)中的任意一种或者多种。 如果没有特别说明, 所述地衣素为选自如下的 7种同系物即式 (I)一 (VII)中的任意一种 或者多种:
(iv)
l-Λ零- -O-Lcy
(VII) 上述式 (I)— (VII)的质荷比 ( ni/z )依次为 993、 1007、 1021、 1035、 1049、 1007、 1021。 所述饮料酒风味化合物包括香气化合物和异嗅化合物, 如果没有特别说明, 所述 香气化合物是指酯类、 醇类和酸类化合物中的任意一种或者多种, 所述异嗅化合物是 指苯酚类化合物中的任意一种或者多种。 具体地, 所述的酯类、 醇类和酸类化合物或 者苯酚类化合物为饮料酒特别是中国饮料酒中的酯类、 醇类和酸类化合物或者苯酚类 化合物。
本发明中, 所述食品优选为饮品; 具体地所述饮品为酒类 (中国白酒、 黄酒、 葡 萄酒、 啤酒、 清酒、 烧酒、 威士忌、 白兰地、 伏特加、 朗姆、 金酒) 。 更具体地, 所 述酒类为白酒例如中国白酒; 进一步具体地, 所述中国白酒为酱香型、 浓香型、 清香 型、 豉香型或药香型白酒。 发明的有益效果
本发明人采用 HS— SPME研究脂肽类化合物(地衣素)对饮料酒中香气化合物的 挥发性影响。 确定在 20— 30 Ό、 30— 50min为萃取温度和萃取时间。 脂肽类化合物特 别是地衣素对各种饮料酒中香气化合物挥发性具有显著的功能作用。 脂肽类化合物特 别是地衣素主要对酒中的酯类、 醇类、 酸类和苯酚类存在着明显的影响。 附图说明
图 1 : 萃取时间对白酒中香气化合物 (酯类、 醇类和酸类) 和异嗅化合物 (苯酚 类) 挥发性的影响。
图 2 : 地衣素的分离收集的洗脱曲线。 (每一个峰对应的是一个同系物) 。
图 3: 地衣素 (质荷比 1035.6 ) 的 MS/MS图谱。
图 4 : 她衣素 (分子量 1034.6 ) 的 1H 核磁图谱 (图 4A)和 13C核磁图谱 (图 4B)。 具体实施方式
下面将结合实施例对本发明的实施方案进行详细描述, 但是本领域技术人员将会 理解, 下列实施例仅用于说明本发明, 而不应视为限定本发明的范围。 实施例中未注 明具体条件者, 按照常规条件或制造商建议的条件进行。 所用试剂或仪器未注明生产 厂商者, 均为可以通过市购获得的常规产品。 删 1: 驗麵》 是舶
1. 实验用品
地衣素 (式 I一式 VI I混合物): 可以商购或者根据现有技术中公开的文献中的记 载进行制备, 也可以参考下面的实施例 4中的方法制备。 本实施例具体使用按照实施 例 4制备的地衣素, 其为七种同系物的混合物 (七种同系物等比例混合) 。
式 V 地衣素: 商购或者参考现有技术例如如下文献制备: Yakimov, M. M.; Abraham, W.-R.; Meyer, H.; Giuliano, L.; Golyshin, P. N., Structural characterization of lichenysin A components by fast atom bombardment tandem mass spectrometry. Biochimica et Biophysica Acta 1999, 1438 (2), 273-280。
式 VII 地衣素: 商购或者参考现有技术例如如下文献制备: Yakimov, M. M.; Abraham, W.-R.; Meyer, H.; Giuliano, L.; Golyshin, P. N., Structural characterization of lichenysin A components by fast atom bombardment tandem mass spectrometry. Biochimica et Biophysica Acta 1999, 1438 (2), 273-280。
白酒酒样: 郎酒酒样 (添加之前, 该酒样中地衣素本身含量 100 g/L; 添加到 1000
2. 实验方法
( 1 )将地衣素应用于酒样中, 混合物地衣素、 式 V地衣素、 式 VI I地衣素的添加 量分别均为 900 Mg/L (即添加到 1000 Mg/L) 。
(2 ) 采用感官评价差异。
感官品评方法如下:
酒样品评由 20位国家白酒评委在室温进行品评打分。
酒样样品稀释酒度至相同的度数 (例如 55度, 不足 55度的可以不稀释) 进行品 评。 感官评价满分 110分, 从色、 香、 味、 风格和空杯留香五个方面进行打分品评, 比较酒质差异, 每个方面的满分依次为 10分、 25分、 50分、 15分、 10分; 其中对于 空杯香, 倒干后放置 30分钟进行打分比较强度。
3. 实验结果
如下面的表 1所示。
表 1: 添加地衣素对酒中感官评价的影响
表 1中的数据显示, 地衣素处理酒样, 白酒综合品质得到显著改善, 空杯 到明显增强。 实施例 2: 模拟体系中 SPME萃取条件的优化
采用 HS-SPME-GC-MS研究几种香气化合物(酯类、 醇类和酸类)和异嗅化合物 (苯酚类)在模拟体系中的挥发性, 首先确定 HS— SPME的实验方法和条件参数, 如 下-
20 mL顶空瓶中加入 8 mL白酒样品, 进行顶空固相微萃取。 顶空固相微萃取的
条件为: 三相萃取头 (DV B/CAR/PDMS, 50/30 μηι ) , 20— 30 °C预热 5 min, 萃取吸 附 30— 50 min, GC解吸 5 min。
GC条件色谱柱: DB-Wax (60 mxO.325 mmx0.25 μιη) ; 升温程序: 50 °C保持 2 min, 以 6 °C/min的速率升至 230 °C, 保持 15 min; 进样口温度 250 °C, 载气 He, 流 速 2 niL/miii; 不分流。
MS条件 EI电离源,离子源温度 230 V,电子能量 70 eV,扫描范围 35.00— 350.00 以几种香气化合物和异嗅化合物的挥发性面积为基准, 确定在 20— 30°C、 30- SOmiii为萃取温度和萃取时间 (见图 1) 。 实施例 3: 地衣素对白酒中的香气化合物的挥发性的影响
1. 实验用品
地衣素 (式 I一式 VII混合物) : 同实施例 1。
式 V地衣素: 同实施例 1。
式 VII地衣素: 同实施例 1。
白酒酒样 1一 3: 分别为郎酒酒样、 浓香酒样、 清香酒样。
2. 实验方法
(1)将地衣素应用于酒样中, 混合物地衣素、 式 V地衣素、 式 VII地衣素的添加 量分别均为 100 g/L。
(2) 影响的评价 /分析参考实施例 2中的方法进行。
3. 实验结果
结果如下面的表 2所示 (空白对照为不添加地衣素) 。
表 2: 添加地衣素对酒中香气化合物和异嗅化合物挥发性的影响
由表 2可见, 地衣素主要对酒中的香气化合物如酯类、 醇类、 酸类, 以及异嗅化 合物苯酚类的挥发性存在着明显的功能作用。 地衣素抑制了苯酚类的挥发, 促进了酯 类、 醇类、 酸类的挥发。 其中, 异嗅化合物苯酚类化合物的挥发明显降低, 即地衣素 有利于降低白酒中异嗅化合物的挥发量, 降低量在 50 %— 70 %之间。 实施例 4: 地衣 ¾对葡 酒、 啤酒中的香气化合物的挥发性的影响
1. 实验用品
地衣素 (式 I一式 VII混合物) : 同实施例 1。
式 V地衣素: 同实施例 1。
式 VI I地衣素: 同实施例 1。
酒样: 葡萄酒、 啤酒。
2. 实验方法
( 1 )将地衣素应用于酒样中, 混合物地衣素、 式 V地衣素、 式 VII地衣素的添加 量分别均为 100 g/L。
( 2 ) 影响的评价 /分析参考实施例 2中的方法进行。
3. 实验结果
结果如下面的表 3所示 (空白对照为不添加地衣素) 。 表 3添加地衣素对葡萄酒、 啤酒中典型异嗅化合物挥发性的影响
cone. 空 白 对添加地衣净 变
Compound odor descriptor source
( δ L) 照 素 化
Phenolic 218681.7 33.19
4-Ethylphenol 440 in wine 146110.58
off-flavor 7 %
4-Ethylguaiaco Phenolic 236798.3 112161.976 52.63
135 in wine
1 off-flavor 3 7 % orris, fat and 200870.2 51.87 trans-2-nonena 20,000 in beer 96688.35
cucumber 6 % 1
104482.0 48527.1233 53.55
Diacetyl 100,000 buttery in beer
4 3 %
Linoleic Acid 100,000 fatty in beer 275077.9 126608.973 53.97
4 3 %
实½例5: 白酒中脂肽类化合 ¾l (地衣素) 的分离纯化
1. 实验仪器和设备
白酒中脂肽类化合物的纯化采用 SPE C18 小柱(Waters),提取仪器为 waters 2767 Sample Manager, Waters 2489 UV/visible Detector , 以及 Waters 2535 Quaternary Gradient Module。
白酒中地衣素的质谱鉴定采用 aters Acquity UPLC和 Synapt Q-TOF system 。
2. 实验方法和实验步骤
(1) 白酒中脂肽类化合物的分离纯化
白酒中挥发性成分的研究较多, 但对其非挥发性成分-大分子化合物的研究相对较 少。 酒样经过提取纯化后的样品, 采用制备液相进行纯化。
酱香型白酒 (郎酒) 经过 SPE C18小柱净化后的样品, 采用制备液相进行纯化。 色谱柱采用 Xbridge Prep C18 (250 mm x 19 mm, 10 μιη)反相色谱柱, 210 nm检测。 分离釆用的流动相配比为: 0.1%的甲酸甲醇溶液 (A) , 甲醇 (B) 。 一次分离程序: 0-10 min, 40%B— 25%B; 10— 50 min, 25%B— 5%B; 50— 55 min, 5%B— 0%B。 样品经过蒸发浓缩后得到纯品。
(2) 结构验证
质谱检测器(MSD) 参数: 正离子模式, 雾化气压 30 psi; 分离电压 3.5 KV, 诱 导碰撞电压 20、,, 干燥气体 N2, 干燥温度 100 V, 离子扫描范围 50— 1500 m/z。 应 用 MaSSlynX 4.1 软件对数据进行采集和分析。
3. 实验结果
从白酒中分离提取纯化得到地衣素。还可以分离出表面素、伊枯草菌素和丰原素。 其中, 地衣素产品分别得到七个同系物化合物。 从图 2可以看出, 经过制备后, 获得 993.6 ( 671.4)、 1007.6 (685.4, 671.4)、 1021.6 (685.4, 671.4)、 1035.6 (685.4)、 1049.6
(685.4)七个不同结构的化合物纯品。
表 4中分别列举了七个纯品的分子量, 母离子, 主要子离子和色谱纯度。
表 4: 地衣素的质谱及纯度
纯度(%) 分子量 M+ (m/z) MS/MS 主要子离子
210 nm
992.6 993.6 671.4 83.7
1006.6 1007.6 685.4 94.8
1006.6 1007.6 671.4 40.6
1020.6 1021.6 685.4 87.3
1020.6 1021.6 671.4 95.3
1034.6 1035.6 685.4 100
1048.6 1049.6 685.4 85.8 实施例 6: 白酒中脂肽类化合物的质谱鉴定
为了确认该化合物的结构信息, 本发明人采用质谱和核磁对其进行化合物鉴定。 采用 Q-TOF-MS对质荷比 1035.6 (685.4)的化合物进行质谱裂解分析。
1. 实验仪器和设备
白酒中脂肽类化合物的质谱鉴定釆用 Waters Acquity UPLC and Synapt Q-TOF system 0
2. 实验方法和实验步骤
采用髙效液相色谱联用质谱进行检测。色谱柱采用 U PLC™ BEH C,s 柱(100 mm x 2.1 mm i.d. , 1.7 μηι ) , 柱温 30。C, 进样体积 2 μί, 流速为 0.3 mL/min。 流动相 A: 0.1 %的甲酸甲醇溶液, B: 甲醇。 洗脱程序: 0— 0.1 min, 15 %B; 0.1— 10 min, 0 % B; 10 - 11 min, 15 % B。 质谱检测器 (MSD ) 参数: 正离子模式, 雾化气压 30 psi; 分离电压 3.5 KV, 诱导碰撞电压 20 V, 干燥气体 N2, 干燥温度 100 °C, 离子扫描范围 50 - 1500 m/z。
3. 实验结果
如图 3所示。
从图 3中可以看出, 其地衣素氨基酸段结构与文献报道的地衣素基本吻合, 氨基 酸段为 N,- Gln-Leu-Leu-Val-Asp-Leii-Ile-C', 脂肪酸段为 C15脂肪酸。
其它 6种同系物化合物的实验数据类似。 实施例 7: 白酒中脂肽类化合物的核磁鉴定
1. 实验仪器和设备
核磁共振仪 (Bruker AM 600)。
2. 实验方法和实验步骤
采用核磁共振仪 (Bruker AM 600) 分析1 H 和 13C NMR图谱, 溶剂为氘代甲醇, 实验温度为 323K。
3. 实验结果
如图 4、 表 5— 6所示。
^ 表 6: 地衣素 (分子量 1034.6) 的 13C核磁图谱化学位移
o o 从图 4 ( A ) 的 1H 谱可以看出, 该化合物中存在一 一 NH— 、 一 ei^KH—和 ο ο
一 一 CH一的化学位移,这些官能团正是形成环肽的重要官能团。其中,一 一 Ο^ΝΗ— ο
存在 7个化学位移和 ― ο CH—存在 1个化学位移, 刚好构成 7个氨基酸与 1个羟基 脂肪酸形成的环肽。 表 4的 Ή谱中的信息比较不全, 不能完全反映化合物的结构信 息。
? I
从图 4 ( Β ) 的 13C 谱可以看出, 该化合物存在着相应的 ―、 一 C'H—G—和
I I
一 CH— NH—的化学位移,这些官能团也正是形成环肽的重要官能团。其中,一 CH— NH— 存在 7个化学位移和一 CH— Q—存在 i个化学位移,刚好构成 7个氨基酸与 1个羟基脂 肪酸形成的环肽。 从核磁数据来看, 该环脂肽必定由 7个氨基酸与 1个羟基脂肪酸形 成的。 表 5的 13C谱中的信息基本反映出化合物的结构信息。
推断地衣素 (分子量 1034.6 ) 的分子式如下:
其它 6种同系物的实验结果类似。 实施例 8: 白酒中脂肽类化合物的分子结构式的确定
经过质谱和核磁数据比对分析, 确认酒中的挥发性一大分子化合物为脂肽类化合 物, 包括表面素、 地衣素、 伊枯草菌素和丰原素。 糊列 8: ά藤口口口巾驗趣
1. 实验仪器和设备
白酒样品中地衣素的定量方法使用 Waters Acquity UPLC 以及 Synapt Q-TOF system 0
2. 实验方法和实验步骤
采用超高效液相色谱联用质谱进行检测。 色谱柱采用 UPLC™ BEH C18柱 (100 mm x 2.1 mm i.d., 1.7 μηι ) , 柱温 30 V, 进样体积 2 μί, 流速为 0.3 mL/min。 流动 相 A: 0.1%的甲酸甲醇溶液, B: 甲醇。 洗脱程序: 0— 0.1 min, 15 B; 0.1-10 min, 0%B; 10-11 min, 15% Bo质谱检测器(MSD)参数: 正离子模式, 雾化气压 30 psi; 分离电压 3.5 KV, 诱导碰撞电压 20 V, 干燥气体 N2, 干燥温度 100Ό, 离子扫描范 围 50— 1500 m/zo
从表 6中的标准曲线计算出其酒中的含量。
3. 实验结果
如表 7— 8所示。
Z0.080/M0ZN3/X3d £Z8丽 SIOZ OAV
从表 7可以看出, 检测限基本在 1 g/L以上, 极大的提高了地衣素的检测灵敏度。 现有的二极管阵列检测手段一般在 10 mg/L以上的检测浓度。
从表 8中可以看出, 白酒中地衣素的含量在 10— lOOO g/L之间, 但是不同的样品之 间地衣素含量差别极大。
结果还表明, 本发明建立了一套高灵敏度的饮品特别是白酒中地衣素的定量测定方 法 (LC— MS/MS) 。
Claims
1. 一种改善或提高饮品香气的方法, 包括使用一种或者多种有效量的脂肽类化合 物的步骤。
2. 脂肽类化合物在制备或生产饮品、 改善或提高饮品香气、 提高饮品中香气化合 物挥发性和 /或降低异嗅化合物挥发性的用途中的用途。
3.根据权利要求 2所述的用途, 其中, 所述香气化合物为酯类、 醇类和酸类化合 物中的任意一种或者多种, 所述异嗅化合物为苯酚类化合物中的任意一种或者多种。
4. 一种白酒生产工艺, 包括使用一种或者多种脂肽类化合物的歩骤。
5. 一种制备脂肽类化合物的方法, 其特征在于使用白酒进行分离纯化;
具体地,
将经过 SPE C18小柱净化后的白酒, 采用制备液相进行纯化;
更具体地,
色谱柱釆用 Xbridge Prep C18 (250 mm x 19 mm, 10 μιη) 反相色谱柱, 210 nm 检测; 分离采用的流动相配比为: 0.1%的甲酸甲醇溶液 (A) , 甲醇(B) ; 一次分离 程序: 0— 10min, 40%B— 25%B; 10— 50 min, 25 B-5 B; 50— 55 min, 5%B—
0%B。
6. 一种测定白酒中脂肽类化合物含量的方法, 其为超高效液相色谱与质谱联用方 法;
具体地,
超高效液相色谱参数: 色谱柱釆用 UPLC™ BEH C18 柱 (100mm x2.1 mmi.d., 1.7 μηι ) , 柱温 30 °C, 进样体积 2 μί, 流速为 0.3 mL/min, 流动相 A: 0.1%的甲酸 甲醇溶液, B: 甲醇; 洗脱程序: 0— O.l min, 15 B; 0.1— lOmin, 0%B; 10— 11 min, 15%B;
质谱检测器 (MSD) 参数: 正离子模式, 雾化气压 30 psi; 分离电压 3.5 KV, 诱
导碰撞电压 20 V, 干燥气体 N2, 干燥温度 100 Ό , 离子扫描范围 50— 1500 m/z。
7. —种组合物, 其含有一种或者多种脂肽类化合物; 可选地, 所述组合物还含有 药学上或者食品学上可接受的载体或者辅料; 具体地, 所述组合物为食品特别是饮品。
8. 根据权利要求 1或 5或 6所述的方法、 权利要求 4所述的白酒生产工艺、 权利 要求 2或 3所述的用途、 或者权利要求 7所述的组合物, 其中,
所述饮品为酒类; 具体地, 所述酒类为白酒例如中国白酒; 更具体地, 所述中国 白酒为酱香型、 浓香型、 清香型、 豉香型或药香型白酒;
所述脂肽类化合物为选自表面素、 地衣素、 伊枯草菌素和丰原素中的任意一种或 者多种;
所述地衣素为选自如下的式 (I)_(VII)中的任意一种或者多种:
-Leu— L-Lcu― i^ciii— CX -
(V)
(VI), 和
(VII)
9. 根据权利要求 1所述的方法、 权利要求 2或 3所述的用途、 权利要求 4所述 的白酒生产工艺、 或者权利要求 7所述的组合物, 其中, 脂肽类化合物的总的含量为 500— 1500 g/L, 优选为 600— 1400 g/L、 700— 1300 g/L、 800— 1200 g/L、 900— 1100 ug/L, 特别优选为 950-1050 ug/L , 例如 1000 g/L。
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