CN110879269A - Combined identification method of cranberry extract - Google Patents
Combined identification method of cranberry extract Download PDFInfo
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Abstract
The invention discloses a combined identification method of cranberry extracts, belonging to the technical field of cranberry extract analysis and identification2Procyanidin B2The method comprises the steps of performing primary identification on a test sample by using a thin-layer analysis method, verifying the specificity of a characteristic peak of cranberry anthocyanin by using a high performance liquid chromatography, comparing the characteristic peak in the test sample with the characteristic peak of the cranberry anthocyanin, and performing secondary identification, and aims to provide the method for identifying the cranberry extract, which has the advantages of sensitive reaction, short detection time and accurate detection result, and accurately judging whether anthocyanin and procyanidin in the cranberry extract are adulterated or not by using a unique anthocyanin spectrogram of the cranberry.
Description
Technical Field
The invention relates to the technical field of cranberry extract analysis and identification, in particular to a combined identification method of cranberry extracts.
Background
Research shows that the cranberry contains a large amount of substances such as vitamin C, iron, an antioxidant, procyanidin (type A), anthocyanin and the like, wherein the procyanidin is also called proanthocyanidin or condensed tannin and belongs to polyphenol substances, bacteria can not be attached to a human body by the procyanidin in the cranberry, so that the probability of acquired infection of the human body is reduced, atherosclerosis can be improved, and cardiovascular diseases are prevented, the anthocyanin in the cranberry belongs to bioflavonoid substances, the physiological activity function of the anthocyanin in the cranberry is represented by free radical scavenging and antioxidation, the most effective antioxidant and the most effective free radical scavenger are found by human beings at present, anthocyanin is formed when the anthocyanin and sugar are combined in the form of a sugar bond, and the anthocyanin in the cranberry exists in the form of anthocyanin.
At present, cranberry extracts are prepared into various foods, beverages and health care products in our markets, but the cranberries in our country mainly depend on import, so the phenomenon of adulteration of the cranberries is more and more severe due to high price, and adulteration modes are various and are too defensive. The cranberry extract adulteration is mainly to add other types of anthocyanidins or procyanidins into the cranberry extract, or other procyanidin-rich and cheaper plant extracts, such as anthocyanidins extracted from grapes, peanuts, pines, mulberry trees, black beans or non-fruit parts of cranberry plants, but the anthocyanidins and procyanidins of the substances have no efficacy contained in the cranberry, and the adulteration addition phenomenon becomes a potential risk of the cranberry extract adulteration. Due to the interference of other impurities in cranberry extracts such as catechin, hippuric acid, vaccinal substance and the like and the influence of complex polymerization molecular structure of procyanidine, other similar characteristic spectrum detection methods such as ' T/CCCHMPIE 1.19-2016 plant extract grape seed extract (grape seed oligomeric procyanidine) ' DB64/T1578-2018 determination high performance liquid chromatography of anthocyanin content in Lycium ruthenicum Murr ' and the like are not applicable to the identification of anthocyanin and procyanidine in cranberry extracts, and because the characteristics of procyanidine spectrum and anthocyanin spectrum in cranberry are not obvious, the sensitivity of contrast substances is extremely low, the currently researched detection methods mostly concentrate on the research of detection of anthocyanin and procyanidine from single sources, for example, the sensitivity is only slightly improved by adopting a fluorescence detector (HPLC-FLD), and the situation that adulteration cannot be identified still exists is likely to be identified, the liquid phase-mass spectrum (HPLC-MS) combination still has the condition that multi-source anthocyanin and procyanidine cannot be distinguished, and the content proportion of quinic acid and malic acid is also adopted to calculate whether cranberry products are adulterated, but because the acids are not unique to cranberries, the detection result is not accurate, and the possibility of adulteration cannot be eliminated, so that a method capable of accurately distinguishing whether other sources of anthocyanin and anthocyanin are adulterated in cranberry extracts is needed at present.
Disclosure of Invention
Aiming at the defects of the prior art, the invention aims to provide the cranberry extract identification method which is sensitive in reaction, short in detection time and accurate in detection result, and the method can be used for accurately judging whether a cranberry extract test sample is adulterated or not by utilizing a unique anthocyanin spectrogram of cranberries.
The invention solves the technical problems by the following technical means:
a method for identifying cranberry extract composition comprises adding catechin and procyanidin A2Procyanidin B2And as a reference, performing primary identification on a test sample by adopting a thin-layer analysis method, verifying the specificity of a characteristic peak of the cranberry anthocyanin by adopting a high performance liquid chromatography, and comparing the characteristic peak in the test sample with the characteristic peak of the cranberry anthocyanin for secondary identification.
Identifying polyphenol procyanidin in the sample by thin layer analysis to identify the added catechin and procyanidin B2As in the case of adulteration, again by highAnd identifying flavonoid anthocyanin in the test sample by using the performance liquid chromatography, and identifying whether a potential counterfeit product is added or not according to the comparison result of the characteristic peak of the cranberry and the characteristic peak of the test sample, thereby further improving the detection accuracy.
Further, the combination authentication method includes the steps of:
A. preliminary identification of whether cranberry extract is adulterated
(A1) Preparing a test article into a solution as a test article A solution;
(A2) preparing a reference substance A solution: respectively collecting catechin and procyanidin A2Procyanidin B2Adding methanol to the standard as reference to obtain mixed solution A, wherein the concentrations of the reference in the mixed solution are 1mg/L respectively;
(A3) thin-layer chromatography analysis: respectively sucking the reference substance A solution and the sample A solution in equal amount, respectively, dropping on the thin layer plate with methanol as blank control, developing with dichloromethane-ethyl acetate-formic acid at volume ratio of 6:10:1 for 15min, taking out, spraying 1% vanillin-hydrochloric acid solution uniformly for coloring, heating at 110 deg.C for 2min, and observing under visible light;
(A4) and (3) chromatographic detection judgment: comparing to determine whether the chromatogram of the sample solution has procyanidin A similar to the chromatogram of the control solution A2Red spots with the same positions, and catechin and procyanidin B in chromatogram of control and comparison test solution2The spot color on the spot and catechin and procyanidin B in the chromatogram of the solution of control A2The spot color on the position is used for judging whether the sample is adulterated or not;
B. performing secondary identification on the solution A of the test article which is identified as not adulterated in the step A;
(B1) selecting a reference medicinal material B:
taking 250mg of freeze-dried fruit powder, adding 20mL of 2% hydrochloric acid-methanol solution, refluxing in hot water bath at 80 ℃ for 10min, cooling, transferring the freeze-dried fruit powder into a 50mL volumetric flask by using the 2% hydrochloric acid-methanol solution, fixing the volume, and filtering by using a 0.45 mu m microporous membrane to obtain a freeze-dried fruit powder solution;
selecting undetermined reference medicinal materials, respectively weighing the undetermined reference medicinal materials, and preparing a undetermined reference medicinal material solution according to a freeze-dried fruit powder solution preparation method;
accurately absorbing the freeze-dried fruit powder solution and the undetermined control medicinal material solution in equal amount respectively, injecting into a high performance liquid chromatograph, measuring, and recording chromatogram;
9 characteristic peaks appear in a chromatogram of the freeze-dried fruit powder solution, the relative retention time is respectively 0.702, 0.780, 0.816, 0.865, 0.893, 1.000, 1.079, 1.173 and 1.236, the standard deviation of the relative retention time of the characteristic peaks appearing in the chromatogram of the undetermined contrast medicinal material solution and the corresponding characteristic peaks of the freeze-dried fruit powder is within +/-5 percent, and the undetermined contrast medicinal material is used as a contrast medicinal material B;
(B2) preparing a reference medicinal material B solution:
weighing the reference medicinal material B determined in the step (B1), and preparing a reference medicine B solution by adopting the same preparation method in the step (B1);
(B3) potential counterfeit solution preparation:
weighing 250mg of the test sample identified as a genuine sample in the step A, and preparing a potential counterfeit solution by adopting the same preparation method in the step (B2);
(B4) preparation of test article B solution:
weighing 250mg of the sample in the step A, and preparing a sample B solution by adopting the same preparation method in the step (B3);
(B5) and (3) detection: accurately absorbing the solution B of the reference medicinal material, the solution of the potential counterfeit product and the solution A of the test sample in equal amount respectively, injecting the solutions into a high performance liquid chromatograph by taking a 2% hydrochloric acid-methanol solution as blank solutions, measuring, and recording a chromatogram;
(B6) the specificity of the cranberry anthocyanin is verified:
comparing the characteristic peaks in the potential counterfeit solution chromatogram with 9 characteristic peaks in the solution chromatogram of the reference medicinal material B one by one, and verifying the specificity of cranberry anthocyanin in the reference medicinal material B;
(B7) and (3) detection and judgment:
if the relative retention time standard deviation of the characteristic peak in the chromatogram of the solution of the test sample B and the 9 characteristic peaks in the chromatogram of the solution of the reference medicinal material B is within +/-5 percent, the test sample is not adulterated;
if the characteristic peaks in the chromatogram of the solution of the test sample B are compared with 9 characteristic peaks in the chromatogram of the solution of the reference medicinal material B, any one of the peaks is deleted or the standard deviation of the relative retention time of the characteristic peaks in the chromatogram of the solution of the test sample B is more than +/-5%, and the test sample is adulterated.
Further, the method for determining chromatography in step (a4) is specifically as follows:
the chromatogram of the sample solution has the same value as the chromatogram of the control A solution2Red spots at the same position, catechin, procyanidin B2The color of the spots on the positions is not darker than that of catechin and procyanidine B in the chromatogram of the solution of the reference substance A2The spot color on the position is not adulterated with the sample;
the chromatogram of the sample solution does not contain procyanidin A in the chromatogram of the reference A solution2Red spots in the same position, or catechin, procyanidin B2The spot color at the position is darker than catechin and procyanidin B in chromatogram of control A solution2Spot color on the site, sample adulteration.
Further, the conditions of the high performance liquid chromatography in the step B are as follows:
a chromatographic column: octadecyl silica gel column 150mm × 4.6mm, 5 μm column temperature: 25 ℃ mobile phase a: 0.5% phosphoric acid
Mobile phase B: acetonitrile-water-acetic acid-phosphoric acid gradient elution time and mobile phase ratio with a volume ratio of 48.5:50:1: 0.5: 0-1 min: 10% B, 90% A, 1-28 min: 10 → 50% B, 90 → 50% A, 28-32 min: 50 → 75% B, 50 → 25% A, 32-32.1 min: 75 → 10% B, 25 → 90% A, 32.1-40 min: 10% B, 90% A;
detection wavelength: flow rate at 520 nm: 1.0mL/min
The theoretical plate number is not less than 60000 calculated according to chlorinated cyanidin.
Further, the undetermined control drug is selected from one of fresh cranberry fruit, concentrated cranberry juice, dried cranberry fruit and freeze-dried cranberry powder.
Further, the potential counterfeits include a grape seed extract solution, a cowberry fruit extract, a mulberry fruit extract, a black bean extract, a pine bark extract, a blackcurrant extract, and a peanut coat extract.
Further, in the (a3) thin layer chromatography step, the thin layer plate was silica gel GF 254.
Further, the preparation steps of the freeze-dried fruit powder are as follows:
cutting fresh cranberry fruits, adding metal particles with the particle size of 500-600 mu m, uniformly stirring, performing vacuum freeze drying, grinding, adding deionized water until the water content is 15%, performing vortex oscillation at 600r/min for 5min under the condition of nitrogen, performing ultrasonic treatment in an ice water bath for 3-5min, wherein the ultrasonic power is 150-200W, the temperature of ice water is 0-2 ℃, and taking out to obtain an aqueous solution of the cranberry fruits; and repeating the vacuum freeze drying operation and ultrasonic ice-water bath treatment under the nitrogen condition on the obtained cranberry aqueous solution, filtering metal particles to obtain cranberry pomace, and crushing the pomace to 200-250 mu m to obtain freeze-dried powder.
Further, the vacuum freeze drying is carried out by freezing at-40 deg.C for 3h, placing into a freeze dryer, vacuumizing to 80-100pa vacuum degree in the freeze dryer, heating to 10 deg.C, drying for 3h, heating to 15 deg.C, and drying for 2 h.
Further, the metal particles are gold particles, and the mass ratio of the gold particles to the fresh cranberry fruits is 3: 1.
In the process of preparing freeze-dried powder from fresh cranberries, multiple times of vacuum freeze drying can prevent water in crushed cranberries from instantly crystallizing when the temperature is too low, so that insufficient freezing of the crushed cranberries is avoided, and the prepared freeze-dried powder has too high water content and larger particles; the added metal particles conduct heat uniformly and rapidly, and can prevent the excessively low local temperature and uneven heating of the cranberry fruit pieces after being mixed with the cranberry fruit pieces, so that the prepared freeze-dried fruit powder is prevented from being poor in dispersibility and stability, not easy to disperse in a solvent and not easy to store. The metal gold has high inertia and uniform heat conduction, can quickly balance the internal temperature of the cranberry fruit pieces, is easy to remove, and can prevent new impurities from being introduced into solid particles. When the freeze-dried fruit powder prepared by the method is subjected to thin-layer analysis and high performance liquid chromatography, impurities are few, excessive interference factors are avoided, chromatogram deviation is avoided, and the credibility for identifying the authenticity of the cranberry extract is improved.
The invention has the beneficial effects that:
(1) identifying catechin and procyanidine A in test sample by thin layer analysis2Procyanidin B2Analyzing whether catechin or procyanidin B is added2And detecting specific anthocyanin and chromatographic distribution in the test sample by combining high performance liquid chromatography to judge whether the sample is adulterated or not.
(2) And (3) carrying out chromatographic analysis on the potential counterfeit, and comparing the potential counterfeit with a specific cranberry anthocyanin chromatographic characteristic peak to verify the specificity of the cranberry anthocyanin characteristic peak so as to promote the identification result to be more accurate.
(3) The freeze-dried fruit powder has low impurity content and few interference factors, improves the identification accuracy, is easy to disperse in a solvent and is easier to store.
(4) The method has accurate result, sensitive reaction and short detection time, is suitable for identifying the cranberry additives in various products taking the cranberry as the raw material, provides convenience for the identification standard of the real and false of the cranberry extract, and has great significance for further development of the industrialization of the cranberry extract.
Drawings
FIG. 1 is a diagram of the method steps of the combined authentication method of the present invention;
FIG. 2 is a thin layer chromatogram of step A of example 1;
FIG. 3 is a high performance liquid chromatogram of the freeze-dried fruit powder of example 1;
FIG. 4 is a high performance liquid chromatogram of commercial fresh fruit 1 of example 1;
FIG. 5 is a high performance liquid chromatogram of commercial fresh fruit 2 of example 1;
FIG. 6 is a high performance liquid chromatogram of commercial fresh fruit 3 of example 1;
FIG. 7 is a high performance liquid chromatogram of commercial fresh fruit 4 of example 1;
FIG. 8 is a high performance liquid chromatogram of commercial fresh fruit 5 of example 1;
FIG. 9 is a high performance liquid chromatogram of commercially available concentrated juice 6 of example 1;
FIG. 10 is a high performance liquid chromatogram of commercially available dried cranberry fruit 7 of example 1;
FIG. 11 is a high performance liquid chromatogram of lyophilized fruit powder 8 of example 1;
FIG. 12 is a high performance liquid chromatogram of chlorinated cyanidin in example 1;
FIG. 13 is a high performance liquid chromatogram of chlorinated paeoniflorin in example 1;
FIG. 14 is a high performance liquid chromatogram of the freeze-dried fruit powder of example 1;
FIG. 15 is a high performance liquid chromatogram of a sample Ba solution in example 1;
FIG. 16 is a high performance liquid chromatogram of a test Bb solution in example 1;
FIG. 17 is a high performance liquid chromatogram of a grape seed extract of example 1;
FIG. 18 is a high performance liquid chromatogram of a Vaccinium myrtillus extract of example 1;
FIG. 19 is a high performance liquid chromatogram of pine bark extract of example 1;
FIG. 20 is a high performance liquid chromatogram of a mulberry extract of example 1;
FIG. 21 is a high performance liquid chromatogram of blackcurrant extract of example 1;
FIG. 22 is a high performance liquid chromatogram of peanut coat extract of example 1;
FIG. 23 is a high performance liquid chromatogram of the black soybean extract in example 1;
FIG. 24 is an air-white control high performance liquid chromatogram of example 1;
FIG. 25 is a thin layer chromatogram for durability detection of the thin layer plate in example 2;
FIG. 26 is a thin layer chromatogram for detection limit investigation in example 2;
FIG. 27 is a thin layer analysis chromatogram of step A in example 3;
FIG. 28 is a high performance liquid chromatogram of a sample Ba solution in example 3;
FIG. 29 is a high performance liquid chromatogram of a test Bb solution of example 3;
FIG. 30 is a high performance liquid chromatogram of a test solution of Bc in example 3;
FIG. 31 is a high performance liquid chromatogram of a solution of the test article Bd in example 3;
FIG. 32 is a high performance liquid chromatogram of a solution of the test article Be in example 3;
FIG. 33 is a high performance liquid chromatogram of a solution of the test article Bf in example 3;
FIG. 34 is a high performance liquid chromatogram of a Bh solution as a test sample in example 3;
FIG. 35 is a high performance liquid chromatogram of a solution of control drug B in example 3;
FIG. 36 is a high performance liquid chromatogram of an air-white control solution of example 3.
Detailed Description
The present invention will be described in detail below with reference to specific embodiments and fig. 1 to 36:
the freeze-dried fruit powder adopted by the invention is prepared by the following steps:
chopping 10g of fresh cranberry, adding 30g of gold particles with the particle size of 500 mu m, uniformly stirring, freezing at the low temperature of-40 ℃ for 3h, then putting into a freeze dryer, vacuumizing until the vacuum degree in the freeze dryer is 100pa, heating to 10 ℃, drying for 3h, heating to 15 ℃, drying for 2h, grinding, adding deionized water until the water content is 15%, performing vortex oscillation at 600r/min for 5min under the protection of nitrogen, performing ultrasonic ice-water bath treatment for 5min under the conditions that the ultrasonic power is 150W and the ice water temperature is 2 ℃, and taking out to obtain a cranberry aqueous solution; repeating the vacuum freeze drying operation on the obtained cranberry fruit aqueous solution, performing ultrasonic ice water bath treatment under the protection of nitrogen, filtering gold particles to obtain cranberry pomace, and crushing the cranberry pomace to 200 mu m to obtain freeze-dried fruit powder.
The terms used in the present invention define: test article A: cranberry extract to be subjected to authenticity identification;
control a: catechin and procyanidin A2Procyanidin B2A mixture of standards;
freeze-drying fruit powder: fruit powder directly prepared from fresh cranberry;
undetermined reference medicinal materials: true cranberries of various forms and states;
and (4) reference medicinal material B: selecting a reference medicinal material which is most convenient for actual use conditions from the reference medicinal materials to be determined;
potential counterfeit products: a substance containing anthocyanidin, wherein the anthocyanidin has a similar component to cranberry anthocyanidin;
solution of test article B: a, identifying a solution prepared from a sample which is a genuine product;
reference solution: respectively preparing chlorinated cyanidin and chlorinated paeoniflorin standard products.
Example 1: cranberry extract identification 1
A. Preliminary identification of whether cranberry extract is adulterated
Test article A: the PAC 95% cranberry extract (obtained from peak health technologies (lot, H180202), PAC 25% cranberry extract (obtained from peak health technologies (lot, H180101) selected in this example;
control a: catechin (Chinese institute for testing food and drug, 110877-201604) and procyanidin A2(SIGMA-ALORICH, LOT # BCBX5559), procyanidin B2(Chengdu Prie method science and technology development, Inc. lot: PRF 9042803);
the methanol adopted when preparing the solution A of the test sample and the solution A of the reference sample is pure chromatogram, and the methanol adopted in the rest other operations is pure chromatogram;
2% hydrochloric acid-methanol solution (2%, m/V) preparation: accurately measuring 47mL of hydrochloric acid (36%, m/V), adding 800mL of methanol, diluting with methanol to 1000mL, and shaking up to obtain the final product;
1% vanillin-hydrochloric acid solution (1%, W/V) preparation: dissolving 1g vanillin in concentrated hydrochloric acid standard solution, diluting to 100ml volume, and shaking up;
(A1) mixing the PAC 95% cranberry extract with methanol to prepare a sample Aa solution with the concentration of 50mg/ml, and mixing the PAC 25% cranberry extract with methanol to prepare a sample Ab solution with the concentration of 100 mg/ml;
(A2) preparation of control A solution: respectively collecting catechin and procyanidin A2Procyanidin B2Adding methanol to obtain a mixed solution as reference A solution, and adding catechin and procyanidin A into the mixed solution2Procyanidin B2The concentration is 1mg/mL respectively;
(A3) thin-layer chromatography analysis: respectively sucking 5ul of test sample Aa solution, Ab solution and control A solution, respectively spotting on the same silica gel GF254 thin layer plate with methanol as blank control, developing with dichloromethane-ethyl acetate-formic acid at volume ratio of 6:10:1 for 15min, taking out, uniformly spraying 1% vanillin-hydrochloric acid solution for coloring, heating at 110 deg.C for 2min, and inspecting under ultraviolet light to obtain chromatograms shown in FIG. 2;
(A4) and (3) chromatographic detection judgment:
as shown in fig. 2, from left to right, 1: control A solution (catechin, procyanidin A)2Procyanidin B2Mixed solution), 2: blank (methanol), 3: test article Aa solution (95% cranberry extract), 4: test Ab solution (25% cranberry extract);
taking the solution of reference substance A as reference, and performing thin layer chromatogram with catechin and procyanidin from top to bottom2Procyanidin B2；
Blank is negative;
and (3) judging:
the appearance of procyanidine A in chromatogram of test Aa solution and the chromatogram of control A solution2Red spots at the same position, catechin, procyanidin B2The color of the spots on the positions is lighter than that of catechin and procyanidine B in the chromatogram of the solution of the reference substance A2Spot color on location, i.e. PAC 95% cranberry extract test article is not adulterated;
the chromatogram of Ab solution of the test sample shows procyanidine A in the chromatogram of A solution of the control sample2Red spots at the same position, catechin, procyanidin B2The color of the spots on the positions is lighter than that of catechin and procyanidine B in the chromatogram of the solution of the reference substance A2Spot color on site, PAC 25% cranberry extract test article, was not adulterated.
B. Performing secondary identification on the test products Aa and Ab which are identified as not adulterated in the step A
Instruments and devices: adopting a high performance liquid chromatograph (with a UV-VIS detector, LC-2030 high performance liquid chromatograph, Shimadzu corporation, China) Co., Ltd.), a BT125D electronic balance (sensitivity of 0.01mg, Sidoristi scientific apparatus, Beijing, Co., Ltd.);
freeze-drying fruit powder: freeze-dried fruit powder (same as freeze-dried fruit powder in step a);
undetermined reference medicinal materials: the fresh cranberry fruits are marked as commercial fresh fruits 1 (producing Canus), commercial fresh fruits 2 (producing Canus), commercial fresh fruits 3 (producing Canus), commercial fresh fruits 4 (producing Canada), commercial fresh fruits 5 (producing Canada), commercial concentrated cranberry juice (Pacific Coast Fruit Products Ltd, lot number 121317, producing Canada, marked as commercial concentrated juice 6), commercial dried cranberry fruits (producing Canada, marked as commercial dried cranberry fruits 7), and freeze-dried Fruit powders (the freeze-dried Fruit powders in step A are the same and marked as freeze-dried Fruit powder 8);
potential counterfeit products: grape seed extract (Tianjin peak), cowberry fruit extract (Yanglingjian), mulberry extract (Shaanxi forest friend), black bean extract (Shaanxi forest friend), pine bark extract (Shaanxi forest friend), blackcurrant extract (Shaanxi forest friend) and peanut skin extract (Shaanxi forest friend);
reference substance: cyanidin chloride (Goodpasture, DST180628-019), paeoniflorin chloride (BATCH: 22).
High performance liquid chromatography conditions:
a chromatographic column: octadecyl bonded silica gel column (150 mm. times.4.6 mm, 5 μm); column temperature: 25 ℃; mobile phase A: 0.5% phosphoric acid; mobile phase B: acetonitrile-water-acetic acid-phosphoric acid in a volume ratio of 48.5:50:1: 0.5;
gradient elution time and mobile phase ratio: 0-1 min: 10% B, 90% A, 1-28 min: 10-50% of B, 90-50% of A, 28-32 min: 50-75% of B, 50-25% of A, 32-32.1 min: 75-10% of B, 25-90% of A, 32.1-40 min: 10% B, 90% A;
detection wavelength: 520 nm; flow rate: 1.0 mL/min; the theoretical plate number is not less than 60000 calculated according to chlorinated cyanidin.
(B1) Selecting a reference medicinal material B:
taking 250mg of freeze-dried fruit powder, adding 20mL of 2% hydrochloric acid-methanol solution, refluxing in hot water bath at 80 ℃ for 10min, cooling, transferring the freeze-dried fruit powder into a 50mL volumetric flask by using the 2% hydrochloric acid-methanol solution, fixing the volume, and filtering by using a 0.45-micrometer microporous membrane to obtain a freeze-dried fruit powder solution;
weighing 1-5 parts of fresh fruits sold in the market, 72.5 g of dried cranberries sold in the market, 6250 ul of concentrated fruit juice sold in the market and 250mg of freeze-dried fruit powder respectively, and preparing into a solution according to a freeze-dried fruit powder solution preparation method;
precisely sucking the above solutions respectively for 10ul, injecting into high performance liquid chromatograph, measuring, and recording chromatogram; the results obtained are shown in FIGS. 3-11, and the collated data are shown in tables 1 and 2:
TABLE 1
Note: the corresponding peak position is marked with "+" and no peak is marked with "-".
TABLE 2
According to the table 1, the table 2 and the fig. 3-11, 9 cranberry anthocyanin characteristic peaks appear in a freeze-dried fruit powder solution chromatogram, the 9 characteristic peaks are numbered as 1-9 in sequence according to the appearance time sequence, and the average relative retention time of the characteristic peaks from 1 to 9 is respectively 0.702, 0.780, 0.816, 0.865, 0.893, 1.000, 1.079, 1.173 and 1.236;
and (3) judging: the standard deviation of the relative retention time of 9 cranberry anthocyanin characteristic peaks appearing in a market fresh fruit 1, a market fresh fruit 2, a market fresh fruit 3, a market fresh fruit 4, a market fresh fruit 5, a market concentrated fruit juice 6, a market dry cranberry fruit 7 and a freeze-dried fruit powder 8 and 9 cranberry anthocyanin characteristic peaks corresponding to the freeze-dried fruit powder is within +/-5%, so that the fresh cranberry fruit, the market concentrated cranberry fruit juice, the market dry cranberry fruit and the freeze-dried fruit powder can be used as a reference medicinal material B of the detection method, and the freeze-dried fruit powder is used as the reference medicinal material B in the detection according to the actual situation.
(B2) Preparing reference medicinal materials and a reference substance B solution:
weighing 250mg of the freeze-dried fruit powder of the control medicinal material B determined in the step (B1), and preparing a solution of the control medicinal material B by adopting the same preparation method of the freeze-dried fruit powder solution of the step (B1);
weighing 5mg of each of chlorinated cyanidin and chlorinated paeoniflorin, respectively placing in 50mL volumetric flasks, adding methanol for ultrasonic dissolution, and filtering with 0.45 μm microporous membrane to obtain reference solution;
(B3) potential counterfeit solution preparation:
respectively weighing 250mg of potential counterfeit products, and preparing potential counterfeit product solution by the same preparation method of the freeze-dried fruit powder solution in the step (B1);
(B4) preparation of a test solution:
respectively weighing the test samples Aa and Ab250mg in the step A, and preparing a test sample solution Ba (PAC 95% cranberry extract) and a test sample solution Bb (PAC 25% cranberry extract) by the same preparation method of the freeze-dried fruit powder solution in the step (B1);
(B5) and (3) detection: accurately sucking 10ul of reference medicinal material solution B, potential counterfeit solution, sample Ba, Bb solution, and reference solution respectively, injecting 2% hydrochloric acid-methanol solution as blank solution into high performance liquid chromatograph, measuring, and recording chromatogram.
The obtained high performance liquid chromatograms are shown in FIGS. 12-24, and the arrangement data is shown in Table 3:
TABLE 3
Note: the corresponding peak position is marked with "+" and no peak is marked with "-".
(B6) The specificity of the cranberry anthocyanin is verified:
determining the positions of chlorinated cyanidin and chlorinated paeoniflorin by using a reference substance solution chromatogram, calculating and determining the relative retention time of main chromatographic peaks in a reference medicinal material B solution by using the chlorinated cyanidin as a reference S peak, and sequencing according to a peak from a small peak to a large peak from a peak 1 to a peak 9, wherein the peak 6 is a chlorinated cyanidin peak, and the peak 8 is the chlorinated paeoniflorin.
Comparing the potential counterfeit solution chromatogram with the control medicinal material B solution chromatogram, the result is as follows:
chromatogram of grape seed extract solution: deletion of peaks 1, 3-5, 7, 9;
blueberry extract chromatogram: deletion of peaks 3, 7, 9;
chromatogram of pine bark extract: deletion of peaks 1-5, 7-9;
chromatogram of mulberry extract: deletion of peaks 1-5, 7, 9;
blackcurrant extract chromatogram: deletion of peaks No. 5, 9;
chromatogram of peanut coat extract: deletion of peaks 3-5, 7;
black bean extract chromatogram: deletion of peaks 1-5, 8-9;
the characteristic peak of the cranberry anthocyanin in the reference medicinal material B is specified;
(B7) and (3) detection and judgment:
if the relative retention time standard deviation of the characteristic peaks in the chromatogram of the Ba solution of the test sample and the Bb solution of the test sample and the 9 characteristic peaks in the chromatogram of the B solution of the reference medicinal material is within +/-5 percent, the test sample is not adulterated.
And (4) conclusion: and comprehensively judging through the step A and the step B that the test sample is not adulterated, namely PAC 95% of cranberry extract and PAC 25% of cranberry extract are not adulterated.
Example 2: durability and detection limit inspection
(1) Durability examination
And (3) taking the same reference substance A solution and blank control as those in the step A of the example 1, carrying out a thin-layer analysis chromatography experiment in the step A, respectively spotting onto GF254 thin-layer plates of different manufacturers, and verifying the durability of the plates according to the result of a thin-layer chromatogram.
The reference medicinal material A is the freeze-dried fruit powder prepared by the invention. Preparing a reference medicinal material A solution: mixing the freeze-dried fruit powder with methanol to prepare a control medicinal material A solution with the concentration of 200 mg/ml.
Adopts a chemical industry research institute of cigarette Tai city GF254 batch No. 20190719, Qingdao ocean chemical Co Ltd GF254 batch No. 20150612; the results obtained by spotting thin-layer plates from three different manufacturers, GF254, Shanghai Xin Yizhai Instrument works Ltd, are shown in FIG. 25: from left to right, the thin-layer plates of shanghai Xinyi, Qingdao ocean chemical industry and Nicoti chemical industry research institute are respectively arranged, and the spectrogram of each thin-layer plate is blank (methanol), a reference substance A solution (mixed by catechin, procyanidin A2 and procyanidin B2) and a reference medicinal material A solution (200mg/ml freeze-dried fruit powder) from left to right, as can be seen from fig. 25, although the three thin-layer plates are from different manufacturers, the detection results of the thin-layer plates of GF254 type are consistent, and the thin-layer plates can be used in the thin-layer analysis process of the invention.
(2) Inspection limit investigation
Preparing the control solution A with the concentration of 1.0, 0.5, 0.2, 0.1 and 0.05mg/ml according to the preparation method of the control solution A in the step A of the example 1, and carrying out sample application, development and inspection on the Qingdao ocean chemical product GF254 of the same thin-layer plate by the same method in the step A under the same condition as the step (A3) of the example 1, wherein the minimum concentration capable of displaying clearly identifiable spots is taken as a detection limit, and the obtained result is shown in figure 26, wherein the spectrograms of the thin-layer plate are the control solution A with the concentration of 1mg/ml, 0.5mg/ml, 0.2mg/ml, 0.1mg/ml and 0.05mg/ml from the left to the right, and the detection limit of catechin of the Qingdao ocean chemical product A thin-layer plate is 0.05mg/ml, and the procyanidine A GF A is calculated from the figure 262The detection limit is 0.1mg/ml, procyanidin B2The detection limit is 0.2 mg/ml.
Example 3: cranberry extract identification II
And (3) testing the sample: randomly purchasing 10 cranberry extracts from different manufacturers from the market as test supplies, and respectively recording the extracts as a, b, c, d, e, f, g, h, i and j;
A. preliminary identification of whether cranberry extract is adulterated
Test article A: taking the test sample as a test sample to be tested, and respectively marking the test sample as Aa, Ab, Ac, Ad, Ae, Af, Ag, Ah, Ai and Aj;
control a: same as example 1;
the methanol adopted when preparing the solution A of the test sample and the solution A of the reference sample is pure chromatogram, and the methanol adopted in other operations is pure chromatogram;
2% hydrochloric acid-methanol solution (2%, m/V) preparation: accurately measuring 47mL of hydrochloric acid (36%, m/V), adding 800mL of methanol, diluting with methanol to 1000mL, and shaking up to obtain the final product;
1% vanillin-hydrochloric acid solution (1%, W/V) preparation: dissolving 1g vanillin in concentrated hydrochloric acid standard solution, diluting to 100ml volume, and shaking up;
(A1) preparing the test articles Aa and Ab into solutions according to the method for preparing the test article Aa solution in the embodiment 1, recording the solutions as the test article Aa and Ab solutions, preparing the test articles Ac-Aj into a solution according to the method for preparing the test article Ab solution in the embodiment 1, and recording the solution as the test article Ac-Aj solution;
(A2) preparing a reference substance A solution: the same procedure as used in example 1 to prepare a solution of control A;
(A3) thin-layer chromatography analysis: respectively sucking 5ul of the test solution Aa-Aj and the control solution A, respectively dropping on the same silica gel GF254 thin layer plate, developing with dichloromethane-ethyl acetate-formic acid at volume ratio of 6:10:1 for 15min, taking out, uniformly spraying 1% vanillin-hydrochloric acid solution for coloring, heating at 110 deg.C for 2min, and observing under ultraviolet light to obtain chromatogram shown in FIG. 27;
(A4) and (3) chromatographic detection judgment:
as shown in fig. 27, from left to right, 1: control A solution (mixed solution of catechin, procyanidin A2 and procyanidin B2), 2-11: a test article Ac-Aj solution;
taking the solution of reference substance A as reference, and the chromatogram map comprises catechin and procyanidin A from top to bottom2Procyanidin B2；
And (3) judging:
the chromatogram of the solution of Aa-Af and Ah of the sample is compared with the chromatogram of the solution of the reference substance A and the chromatogram of the solution of the reference medicinal material A2Red spots at the same position, catechin, procyanidin B2The color of the spots on the positions is lighter than that of catechin and procyanidine B in the chromatogram of the solution of the reference substance A2Spot color on the position, namely the samples Aa-Af and Ah are not adulterated;
the sample Ag, Ai, Aj solution chromatogram does not appear in comparison with procyanidine A in the reference A solution chromatogram2And (4) the red spots on the positions are the same, namely the samples Ag, Ai and Aj are adulterated.
B. Performing secondary identification on the test products Aa-Af and Ah which are identified as not adulterated in the step A
Instruments and devices: same as example 1;
reference substance: same as in example 1.
High performance liquid chromatography conditions: same as in example 1.
(B1) Selecting a reference medicinal material B: the reference drug B is freeze-dried fruit powder (same as the example 1);
(B2) preparing reference medicinal materials and a reference substance B solution: the preparation method is the same as that of the example 1;
(B3) preparation of test article B solution:
respectively weighing 250mg of the test products Aa-Af and Ah in the step A, and preparing a test product Ba-Bf and Bh solution by adopting the same preparation method as the freeze-dried fruit powder solution in the step (B1) in the example 1; taking a 2% hydrochloric acid-methanol solution as a blank solution;
(B4) and (3) detection: accurately sucking 10ul of reference medicinal material B solution, sample Ba-Bf, Bh solution, blank reference solution, and reference solution respectively, injecting into high performance liquid chromatograph, measuring, and recording chromatogram.
The obtained high performance liquid chromatograms are shown in FIGS. 28 to 36, and the arrangement data is shown in Table 4:
TABLE 4
Note: the corresponding peak position is marked with "+" and no peak is marked with "-".
(B5) The specificity of the cranberry anthocyanin is verified:
the same method as that in example 1 is adopted to verify that the cranberry anthocyanin in the reference medicinal material B has specificity;
(B6) and (3) detection and judgment:
if the relative retention time standard deviation of the characteristic peaks in the solution chromatograms of the test sample Ba-Bf and Bh and the 9 characteristic peaks in the solution chromatogram of the reference medicinal material B is within +/-5 percent, the test sample is not adulterated;
if the characteristic peaks in the chromatogram of the Ba-Bf and Bh solutions of the test sample are compared with the 9 characteristic peaks in the chromatogram of the B solution of the reference medicinal material, the standard deviation of the relative retention time of any one of the missing peaks or the characteristic peaks in the chromatogram of the B solution of the test sample is more than +/-5%, and the test sample is adulterated.
The standard deviation of the relative retention time of the characteristic peak in the high performance liquid chromatogram of the Ba-Be solution of the test sample and the 9 characteristic peaks in the chromatogram of the solution of the reference medicinal material B is within +/-5 percent, namely the Ba-Be solution of the test sample is not adulterated.
And (3) comparing the characteristic peaks in the high performance liquid chromatogram of the test sample Bf solution with the 9 characteristic peaks in the chromatogram of the reference medicinal material B solution, and deleting the No. 3 and No. 5 peaks, namely, the test sample Bf is adulterated.
Compared with 9 characteristic peaks in the chromatogram of the solution of the reference medicinal material B, the characteristic peaks in the high-performance liquid chromatogram of the solution of the sample Bh lack peaks 1, 2, 4 and 5, and the sample Bh is adulterated.
And (4) conclusion: and comprehensively judging through the step A and the step B, wherein the samples a, B, c, d and e are not adulterated, and the samples f, g, h, i and j are adulterated.
Although the present invention has been described in detail with reference to the preferred embodiments, it will be understood by those skilled in the art that various changes may be made and equivalents may be substituted without departing from the spirit and scope of the invention as defined in the appended claims. The techniques, shapes, and configurations not described in detail in the present invention are all known techniques.
Claims (10)
1. A method for identifying cranberry extract composition comprises the steps of combining catechin and procyanidin A2Procyanidin B2For reference, the sample is first identified by thin layer analysisAnd (3) verifying the specificity of the characteristic peak of the cranberry anthocyanin by using the high performance liquid chromatography, and comparing the characteristic peak in the test sample with the characteristic peak of the cranberry anthocyanin for secondary identification.
2. The method of identifying a combination of cranberry extracts of claim 1, comprising the steps of:
A. preliminary identification of whether cranberry extract is adulterated
(A1) Preparing a test article into a solution as a test article A solution;
(A2) preparing a reference substance A solution: respectively collecting catechin and procyanidin A2Procyanidin B2Adding methanol as reference substance to obtain mixed solution A, wherein the concentrations of the reference substances in the mixed solution are 1mg/L respectively;
(A3) thin-layer chromatography analysis: respectively sucking a reference substance A solution and a test substance A solution in an equal amount, respectively, taking methanol as a blank reference, respectively dropping on a thin-layer plate, developing for 15min by using a dichloromethane-ethyl acetate-formic acid mixed solution with a volume ratio of 6:10:1, taking out, uniformly spraying 1% vanillin-hydrochloric acid solution for coloring, heating at 110 ℃ for 2min, and observing under visible light;
(A4) and (3) chromatographic detection judgment: comparison judgment
Whether the chromatogram of the test solution has procyanidin A in the chromatogram of the reference solution A2Red spots with the same positions, and catechin and procyanidin B in chromatogram of control and comparison test solution2The spot color on the spot and catechin and procyanidin B in the chromatogram of the solution of control A2The spot color on the position is used for judging whether the sample is adulterated or not;
B. performing secondary identification on the sample which is identified as not adulterated in the step A;
(B1) selecting a reference medicinal material B:
taking 250mg of freeze-dried fruit powder, adding 20mL of 2% hydrochloric acid-methanol solution, refluxing in hot water bath at 80 ℃ for 10min, cooling, transferring the freeze-dried fruit powder into a 50mL volumetric flask by using the 2% hydrochloric acid-methanol solution, fixing the volume, and filtering by using a 0.45 mu m microporous membrane to obtain a freeze-dried fruit powder solution;
selecting undetermined reference medicinal materials, respectively weighing the undetermined reference medicinal materials, and preparing a undetermined reference medicinal material solution according to a freeze-dried fruit powder solution preparation method;
accurately absorbing the freeze-dried fruit powder solution and the undetermined control medicinal material solution in equal amount respectively, injecting into a high performance liquid chromatograph, measuring, and recording chromatogram;
9 characteristic peaks appear in a chromatogram of the freeze-dried fruit powder solution, the relative retention time is respectively 0.702, 0.780, 0.816, 0.865, 0.893, 1.000, 1.079, 1.173 and 1.236, the standard deviation of the relative retention time of the characteristic peaks appearing in the chromatogram of the undetermined contrast medicinal material solution and the corresponding characteristic peaks of the freeze-dried fruit powder is within +/-5 percent, and the undetermined contrast medicinal material is used as a contrast medicinal material B;
(B2) preparing a reference medicinal material B solution:
weighing the reference medicinal material B determined in the step (B1), and preparing a reference medicine B solution by adopting the same preparation method in the step (B1);
(B3) potential counterfeit solution preparation:
respectively weighing 250mg of potential counterfeit products, and preparing a potential counterfeit product solution by the same preparation method in the step (B2);
(B4) preparation of test article B solution:
weighing 250mg of the test sample identified as a genuine sample in the step A, and preparing a test sample B solution by adopting the same preparation method in the step (B3);
(B5) and (3) detection: accurately absorbing the reference medicinal material solution B, the potential counterfeit solution and the sample solution B in equal amount, respectively, injecting the blank solutions into a high performance liquid chromatograph by using 2% hydrochloric acid-methanol solution, measuring, and recording chromatogram;
(B6) the specificity of the cranberry anthocyanin is verified:
comparing the characteristic peaks in the potential counterfeit solution chromatogram with 9 characteristic peaks in the solution chromatogram of the reference medicinal material B one by one, and verifying the specificity of cranberry anthocyanin in the reference medicinal material B;
(B7) and (3) detection and judgment:
if the relative retention time standard deviation of the characteristic peak in the chromatogram of the solution of the test sample B and the 9 characteristic peaks in the chromatogram of the solution of the reference medicinal material B is within +/-5 percent, the test sample is not adulterated;
if the characteristic peaks in the chromatogram of the solution of the test sample B are compared with 9 characteristic peaks in the chromatogram of the solution of the reference medicinal material B, any one of the peaks is deleted or the standard deviation of the relative retention time of the characteristic peaks in the chromatogram of the solution of the test sample B is more than +/-5%, and the test sample is adulterated.
3. The combination identification method of cranberry extracts according to claim 2, wherein the chromatographic detection and judgment method in step (a4) is as follows:
the chromatogram of the sample solution has the same value as the chromatogram of the control A solution2Red spots at the same position, catechin, procyanidin B2The color of the spots on the positions is not darker than that of catechin and procyanidine B in the chromatogram of the solution of the reference substance A2The spot color on the position is not adulterated with the sample;
the chromatogram of the sample solution does not contain procyanidin A in the chromatogram of the reference A solution2Red spots in the same position, or catechin, procyanidin B2The spot color at the position is darker than catechin and procyanidin B in chromatogram of control A solution2Spot color on the site, sample adulteration.
4. The method for identifying the combination of cranberry extracts of claim 3, wherein the HPLC conditions in step B are as follows:
a chromatographic column: octadecyl bonding silica gel column 150mm × 4.6mm, 5 μm
Column temperature: 25 deg.C
Mobile phase A: 0.5% phosphoric acid
Mobile phase B: acetonitrile-water-acetic acid-phosphoric acid gradient elution time and mobile phase ratio with a volume ratio of 48.5:50:1: 0.5: 0-1 min: 10% B, 90% A, 1-28 min: 10 → 50% B, 90 → 50% A, 28-32 min: 50 → 75% B, 50 → 25% A, 32-32.1 min: 75 → 10% B, 25 → 90% A, 32.1-40 min: 10% B, 90% A;
detection wavelength: 520nm
Flow rate: 1.0mL/min
The theoretical plate number is not less than 60000 calculated according to chlorinated cyanidin.
5. The method of claim 4, wherein the candidate control drug is selected from one of fresh cranberry fruit, concentrated cranberry juice, dried cranberry fruit, and lyophilized cranberry powder.
6. The method of identifying a combination of cranberry extracts of claim 5, wherein the potential counterfeit comprises grape seed extract, blueberry extract, mulberry extract, black bean extract, pine bark extract, blackcurrant extract, peanut coat extract.
7. The combination identification method for cranberry extracts according to claim 6, wherein in the step of (A3) thin layer chromatography, the thin layer plate is silica gel GF 254.
8. The combination identification method of cranberry extracts of claim 7, wherein the freeze-dried fruit powder is prepared by the following steps:
cutting fresh cranberry fruits, adding metal particles with the particle size of 500-600 mu m, uniformly stirring, performing vacuum freeze drying, grinding, adding deionized water until the water content is 15%, performing vortex oscillation at 600r/min for 5min under the condition of nitrogen, performing ultrasonic treatment in an ice water bath for 3-5min, wherein the ultrasonic power is 200W, the temperature of ice water is 0-2 ℃, and taking out to obtain an aqueous solution of the cranberry fruits; and repeating the vacuum freeze drying operation and ultrasonic ice-water bath treatment under the nitrogen condition on the obtained cranberry aqueous solution, filtering metal particles to obtain cranberry pomace, and crushing the pomace to 200-250 mu m to obtain freeze-dried powder.
9. The method for identifying the combination of cranberry extracts as claimed in claim 8, wherein the vacuum freeze-drying is performed by freezing at-40 ℃ for 3 hours, then placing in a freeze-dryer, vacuumizing to 80-100pa vacuum degree in the freeze-dryer, heating to 10 ℃, drying for 3 hours, heating to 15 ℃, and drying for 2 hours.
10. The method of claim 9, wherein the metal particles are gold particles, and the mass ratio of gold particles to fresh cranberry fruits is 3: 1.
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Citations (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001052671A2 (en) * | 2000-01-24 | 2001-07-26 | Dry Creek Nutrition, Inc. | A method of altering and improving taste characteristics of edible consumables with monomeric or oligomeric polyphenolic compounds |
| CN101113411A (en) * | 2007-06-27 | 2008-01-30 | 东华大学 | Induction preparation of EGCG esterase and method for producing EGC and gallic acid by using the enzyme |
| CN101125844A (en) * | 2007-09-28 | 2008-02-20 | 天津市尖峰天然产物研究开发有限公司 | Method for extracting procyanidins B2 |
| CN101239962A (en) * | 2007-02-07 | 2008-08-13 | 上海华珠生物科技有限公司 | Method for extracting proanthocyanidins from cranberry |
| CN101250206A (en) * | 2008-03-26 | 2008-08-27 | 天津市尖峰天然产物研究开发有限公司 | Effective part containing cyaniding 3-O-glucoside as well as preparation method and use thereof |
| CN101498703A (en) * | 2009-02-12 | 2009-08-05 | 安徽农业大学 | Cell line selection method for tea plant containing different catechin content and components |
| CN102526170A (en) * | 2011-12-31 | 2012-07-04 | 许学志 | Catechu extract composition for resisting tubercle bacillus, preparation method of catechu extract composition, pharmaceutical preparation containing catechu extract composition, and application of catechu extract composition |
| CN104623670A (en) * | 2013-11-06 | 2015-05-20 | 高松 | Compositions Containing Enriched Natural Crocin and/or Crocetin, and Their Therapeutic or Nutraceutical Uses |
| US10583218B2 (en) * | 2008-11-05 | 2020-03-10 | Hancock Jaffe Laboratories Aesthetics, Inc. | Composite containing collagen and elastin as a dermal expander and tissue filler |
-
2019
- 2019-12-10 CN CN201911259149.0A patent/CN110879269B/en active Active
Patent Citations (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001052671A2 (en) * | 2000-01-24 | 2001-07-26 | Dry Creek Nutrition, Inc. | A method of altering and improving taste characteristics of edible consumables with monomeric or oligomeric polyphenolic compounds |
| CN101239962A (en) * | 2007-02-07 | 2008-08-13 | 上海华珠生物科技有限公司 | Method for extracting proanthocyanidins from cranberry |
| CN101113411A (en) * | 2007-06-27 | 2008-01-30 | 东华大学 | Induction preparation of EGCG esterase and method for producing EGC and gallic acid by using the enzyme |
| CN101125844A (en) * | 2007-09-28 | 2008-02-20 | 天津市尖峰天然产物研究开发有限公司 | Method for extracting procyanidins B2 |
| CN101250206A (en) * | 2008-03-26 | 2008-08-27 | 天津市尖峰天然产物研究开发有限公司 | Effective part containing cyaniding 3-O-glucoside as well as preparation method and use thereof |
| US10583218B2 (en) * | 2008-11-05 | 2020-03-10 | Hancock Jaffe Laboratories Aesthetics, Inc. | Composite containing collagen and elastin as a dermal expander and tissue filler |
| CN101498703A (en) * | 2009-02-12 | 2009-08-05 | 安徽农业大学 | Cell line selection method for tea plant containing different catechin content and components |
| CN102526170A (en) * | 2011-12-31 | 2012-07-04 | 许学志 | Catechu extract composition for resisting tubercle bacillus, preparation method of catechu extract composition, pharmaceutical preparation containing catechu extract composition, and application of catechu extract composition |
| CN104623670A (en) * | 2013-11-06 | 2015-05-20 | 高松 | Compositions Containing Enriched Natural Crocin and/or Crocetin, and Their Therapeutic or Nutraceutical Uses |
Non-Patent Citations (4)
| Title |
|---|
| ANNA ENGEMANN 等: "Intestinal Metabolism of Two A-type Procyanidins Using the Pig Cecum Model: Detailed Structure Elucidation of Unknown Catabolites with Fourier Transform Mass Spectrometry (FTMS)", 《JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY》 * |
| RODRIGO P. FELICIANO 等: "Comparison of Isolated Cranberry (Vaccinium macrocarpon Ait.) Proanthocyanidins to Catechin and Procyanidins A2 and B2 for Use as Standards in the 4-(Dimethylamino)cinnamaldehyde Assay", 《JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY》 * |
| VESNA GLAVNIK 等: "Comparison of TLC and HPLC Methods Used for Analysis of (–)-Epicatechin and its Dimer Procyanidin B2 in Chocolate", 《JOURNAL OF PLANAR CHROMATOGRAPHY》 * |
| 姚立君 等: "蔓越莓营养与保健功能研究进展", 《食品研究与开发》 * |
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