CN113030329B - One-test-multiple-evaluation method for determining content of 6 gingerol components in ginger juice brown sugar - Google Patents
One-test-multiple-evaluation method for determining content of 6 gingerol components in ginger juice brown sugar Download PDFInfo
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- JZLXEKNVCWMYHI-UHFFFAOYSA-N gingerol Natural products CCCCC(O)CC(=O)CCC1=CC=C(O)C(OC)=C1 JZLXEKNVCWMYHI-UHFFFAOYSA-N 0.000 title claims abstract 17
- NLDDIKRKFXEWBK-AWEZNQCLSA-N gingerol Chemical compound CCCCC[C@H](O)CC(=O)CCC1=CC=C(O)C(OC)=C1 NLDDIKRKFXEWBK-AWEZNQCLSA-N 0.000 title claims abstract 12
- 241000234314 Zingiber Species 0.000 title claims abstract 10
- 235000006886 Zingiber officinale Nutrition 0.000 title claims abstract 10
- 235000008397 ginger Nutrition 0.000 title claims abstract 10
- 235000011389 fruit/vegetable juice Nutrition 0.000 title claims abstract 9
- 238000011156 evaluation Methods 0.000 title claims abstract 3
- FADFGCOCHHNRHF-VAWYXSNFSA-N [10]-Shogaol Chemical compound CCCCCCCCC\C=C\C(=O)CCC1=CC=C(O)C(OC)=C1 FADFGCOCHHNRHF-VAWYXSNFSA-N 0.000 claims abstract 10
- 238000000034 method Methods 0.000 claims abstract 10
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 claims abstract 9
- 238000001514 detection method Methods 0.000 claims abstract 9
- AIULWNKTYPZYAN-SFHVURJKSA-N (10)-Gingerol Chemical compound CCCCCCCCC[C@H](O)CC(=O)CCC1=CC=C(O)C(OC)=C1 AIULWNKTYPZYAN-SFHVURJKSA-N 0.000 claims abstract 5
- OQWKEEOHDMUXEO-UHFFFAOYSA-N (6)-shogaol Natural products CCCCCC=CC(=O)CCC1=CC=C(O)C(OC)=C1 OQWKEEOHDMUXEO-UHFFFAOYSA-N 0.000 claims abstract 5
- BCIWKKMTBRYQJU-INIZCTEOSA-N (8)-Gingerol Chemical compound CCCCCCC[C@H](O)CC(=O)CCC1=CC=C(O)C(OC)=C1 BCIWKKMTBRYQJU-INIZCTEOSA-N 0.000 claims abstract 5
- ZEASWHWETFMWCV-UHFFFAOYSA-N 7-O-(2-O-Acetyl-6-O-Methyl-beta-D-glucuronoside)-4',5,7-Trihydroxyflavone Natural products C=1C(O)=C(O)C2=C(O)C(=O)C=C(C3C(CC4=C(O)C=C(O)C=C4O3)OC(=O)C=3C=C(O)C(O)=C(O)C=3)C=C2C=1C1OC2=CC(O)=CC(O)=C2CC1OC(=O)C1=CC(O)=C(O)C(O)=C1 ZEASWHWETFMWCV-UHFFFAOYSA-N 0.000 claims abstract 5
- VSDUZFOSJDMAFZ-UHFFFAOYSA-N 8-gingerol Natural products COC(=O)C(N)CC1=CC=CC=C1 VSDUZFOSJDMAFZ-UHFFFAOYSA-N 0.000 claims abstract 5
- LGZSMXJRMTYABD-UHFFFAOYSA-N 8-shogaol Natural products CCCCCCCC=CC(=O)CCC1=CC=C(O)C(OC)=C1 LGZSMXJRMTYABD-UHFFFAOYSA-N 0.000 claims abstract 5
- AIULWNKTYPZYAN-UHFFFAOYSA-N 810gingerol Natural products CCCCCCCCCC(O)CC(=O)CCC1=CC=C(O)C(OC)=C1 AIULWNKTYPZYAN-UHFFFAOYSA-N 0.000 claims abstract 5
- OQWKEEOHDMUXEO-BQYQJAHWSA-N [6]-Shogaol Chemical compound CCCCC\C=C\C(=O)CCC1=CC=C(O)C(OC)=C1 OQWKEEOHDMUXEO-BQYQJAHWSA-N 0.000 claims abstract 5
- LGZSMXJRMTYABD-MDZDMXLPSA-N [8]-Shogaol Chemical compound CCCCCCC\C=C\C(=O)CCC1=CC=C(O)C(OC)=C1 LGZSMXJRMTYABD-MDZDMXLPSA-N 0.000 claims abstract 5
- OAHWPNUPCSDOGU-UHFFFAOYSA-N trans-10-shogaol Natural products CCCCCCCCCC=CC(=O)CCc1ccc(O)c(CO)c1 OAHWPNUPCSDOGU-UHFFFAOYSA-N 0.000 claims abstract 5
- 238000010828 elution Methods 0.000 claims abstract 3
- 238000004128 high performance liquid chromatography Methods 0.000 claims abstract 2
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims 6
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 claims 3
- 239000000523 sample Substances 0.000 claims 2
- 239000000243 solution Substances 0.000 claims 2
- 238000001914 filtration Methods 0.000 claims 1
- 238000002347 injection Methods 0.000 claims 1
- 239000007924 injection Substances 0.000 claims 1
- 239000012982 microporous membrane Substances 0.000 claims 1
- RGOVYLWUIBMPGK-UHFFFAOYSA-N nonivamide Chemical compound CCCCCCCCC(=O)NCC1=CC=C(O)C(OC)=C1 RGOVYLWUIBMPGK-UHFFFAOYSA-N 0.000 claims 1
- 239000013558 reference substance Substances 0.000 claims 1
- 239000012488 sample solution Substances 0.000 claims 1
- 238000009210 therapy by ultrasound Methods 0.000 claims 1
- 238000005303 weighing Methods 0.000 claims 1
- YKPUWZUDDOIDPM-SOFGYWHQSA-N capsaicin Chemical compound COC1=CC(CNC(=O)CCCC\C=C\C(C)C)=CC=C1O YKPUWZUDDOIDPM-SOFGYWHQSA-N 0.000 abstract 2
- 229960002504 capsaicin Drugs 0.000 abstract 1
- 235000017663 capsaicin Nutrition 0.000 abstract 1
- 238000003908 quality control method Methods 0.000 abstract 1
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Abstract
The invention provides a one-test and multi-evaluation method for determining the content of 6 gingerol components in ginger juice brown sugar. The method is based on high performance liquid chromatography, ECOSIL C18-eco100A C (4.6X 250mm,5 μm) is used as a chromatographic column, acetonitrile-1% acetic acid solution is used as a mobile phase, gradient elution is carried out, the flow rate is 1.0ml/min, the column temperature is 35 ℃, and the detection wavelength is 280nm. The result shows that in the linear range, the relative correction factors of the synthesized capsaicin, the 6-gingerol, the 8-gingerol, the 10-gingerol, the 6-shogaol, the 8-shogaol and the 10-shogaol are 0.9433, 1.6183, 1.2666, 1.3006, 1.0369 and 1.2355 respectively, the reproducibility of the relative correction factors is good under different experimental conditions, and the calculated values and the measured values of 6 components have no significant difference. Therefore, the method for determining the components of 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol and 10-shogaol in the ginger juice brown sugar is accurate and feasible, and can be used for quality control of the ginger juice brown sugar.
Description
Technical Field
The invention relates to the technical field of analytical chemistry, in particular to a one-test-multiple-evaluation method for determining contents of 6 gingerol components in ginger juice brown sugar.
Background
The ginger juice brown sugar is edible sugar prepared by adding ginger juice and ginger powder into brown sugar serving as a raw material. The traditional Chinese medicine nutriology considers that the brown sugar with warm nature plays a role in enriching blood by 'warming and tonifying, warming and dredging, warming and dispersing', and the ginger has the effects of relieving exterior syndrome and dispelling cold, warming middle-jiao and preventing vomiting, warming lung and relieving cough and detoxifying, and is commonly used for treating cold due to wind-cold, spleen and stomach cold, stomach cold and vomiting, lung cold and cough and relieving fish and crab toxicity. Folk agents have the label of "male can not be without whooping without ginger, female can not be without whooping without sugar". Ginger soup decocted by ginger and brown sugar can activate blood circulation, dispel cold, prevent and treat cold, and is a good dietetic therapy medicine for wind-cold type cold from old times.
At present, a lot of products related to ginger juice brown sugar are available on the market, but no national standard related to the products exists. Although the light industry standard QB/T5006-2016 specifies various indexes of ginger juice brown sugar, the standard is not mandatory, and almost all products in the market are GB/T29602 solid beverages or own enterprise labels. So that we found in the previous investigation that the addition amount of ginger was very different among the products.
The main components of the ginger juice brown sugar are sucrose and gingerol. The number of common gingerol components is 6, and at present, 6-gingerol, 8-gingerol and 10-gingerol are only specified in 2020 pharmacopoeia as index components for quality evaluation of ginger and are not comprehensive enough. In GB/T22293-2008, 6 components are measured by a correction factor method, but the peak positioning is very fuzzy, the separation is not ideal, the interference is large, and the durability and the like are not clear.
However, no literature report that the main active ingredients of the gingerol and the gingerol are used as the quality evaluation indexes of the ginger juice brown sugar is found at present. Therefore, the research establishes a one-test-multiple-evaluation method to simultaneously analyze the contents of gingerol and gingerol of different products, and aims to take the main characteristic active ingredients of ginger, namely 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol and 10-shogaol, as the quality control index ingredients of the ginger, so that the quality of the ginger juice brown sugar can be evaluated more comprehensively, and enterprises and institutions can execute and utilize the ginger juice brown sugar conveniently.
Disclosure of Invention
The invention aims to provide a one-test-and-multiple-evaluation method for determining the content of 6 gingerol components in ginger juice brown sugar, aiming at solving the technical problems that the conventional method is difficult to simultaneously determine six gingerol components of 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol and 10-shogaol and the content of the gingerol components.
Another technical problem to be solved by the present invention is how to further improve the precision, stability, repeatability and durability of the detection method.
In order to achieve the technical purpose, the invention adopts the following technical scheme:
a one-test-multiple-evaluation method for determining the content of 6 gingerol components in ginger juice brown sugar is a high performance liquid chromatography, and comprises the following steps: ECOSIL C18-eco100A C is used as a chromatographic column, acetonitrile-1% acetic acid solution is used as a mobile phase, gradient elution is carried out, the flow rate is 1.0ml/min, the column temperature is 35 ℃, and the detection wavelength is 280nm.
Preferably, the method comprises measuring relative correction factors of six components, namely 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol and 10-shogaol, by using the externally added synthetic capsaicin as a reference substance, and calculating the contents of the six components by using the relative correction factors.
Preferably, the ECOSIL C18-eco100A C column size is 4.6X 250mm,5 μm.
Preferably, the amount of sample used in this method is 20. Mu.l.
Preferably, the mobile phase is a 1% acetic acid solution as component A, acetonitrile as component B, and the gradient elution conditions are as follows: 0-15.0min, 52.0%; 15.0-17.0min, 52.0%; 17.0-43.0min, 40.0% by A → 20.0% by weight A; 43.0-43.01min, 20.0% by weight A → 52.0% by weight A; 43.01-50.0 min,52.0% A.
Preferably, in the method, the sample solution is subjected to the following treatment before sample injection: weighing 1.00g of ginger sugar, dissolving the ginger sugar in methanol, carrying out vortex treatment for 1min, then carrying out ultrasonic treatment for 30min, then using the methanol to fix the volume to 10ml, then shaking up and filtering by a 0.22 mu m microporous membrane.
Preferably, the detection range of six components of 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol and 10-shogaol is 2-50 mu g/ml.
Preferably, the detection limit of 6-gingerol is 5.56mg/kg; the detection limit of 8-gingerol is 8.77mg/kg; the detection limit of 10-gingerol is 7.81mg/kg; the detection limit of 6-shogaol is 7.11mg/kg; the detection limit of 8-shogaol is 9.43mg/kg; the detection limit of 10-shogaol is 8.57mg/kg.
Preferably, in the method, the flow rate fluctuation range is not more than 0.7-1.5 ml/min; the fluctuation range of the column temperature is not more than 30-40 ℃; the fluctuation range of the pH value of the mobile phase is not more than 1.5-3.5.
The invention provides a one-test and multi-evaluation method for determining the content of 6 gingerol components in ginger juice brown sugar. The method is based on high performance liquid chromatography, ECOSIL C18-eco100A C (4.6X 250mm,5 μm) is used as a chromatographic column, acetonitrile-1% acetic acid solution is used as a mobile phase, gradient elution is carried out, the flow rate is 1.0ml/min, the column temperature is 35 ℃, and the detection wavelength is 280nm. Respectively measuring relative correction factors of 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol and 10-shogaol by taking the added synthetic capsaicin as a reference substance, and calculating the contents of the 6 components in the ginger juice brown sugar by using the relative correction factors; meanwhile, the content of 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol and 10-shogaol is determined by an external standard method, and the difference of the two determination methods is compared.
The result shows that in the linear range, the relative correction factors of the synthesized capsaicin, the 6-gingerol, the 8-gingerol, the 10-gingerol, the 6-shogaol, the 8-shogaol and the 10-shogaol are 0.9433, 1.6183, 1.2666, 1.3006, 1.0369 and 1.2355 respectively, the reproducibility of the relative correction factors is good under different experimental conditions, and the calculated values and the measured values of 6 components have no significant difference. Therefore, the method for determining the components of 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol and 10-shogaol in the ginger juice brown sugar is accurate and feasible, and can be used for quality control of the ginger juice brown sugar.
Drawings
FIG. 1 is a graph of the absorption spectra (scanning wavelength 190-640 nm) of 6 gingerol components and synthetic capsaicin; in the figure, 1 to 7 are respectively: 6-gingerol, synthetic capsaicin, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol, 10-shogaol.
FIG. 2 is a chromatogram of a mixed standard solution and an extract of brown sugar from ginger juice; in the figure, A is a mixed reference solution, and B is a sample; 1 to 7 are respectively: 6-gingerol, synthetic capsaicin, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol, 10-shogaol.
Detailed Description
Hereinafter, specific embodiments of the present invention will be described in detail. Well-known structures or functions may not be described in detail in the following embodiments in order to avoid unnecessarily obscuring the details. Approximating language, as used herein in the following examples, may be applied to identify quantitative representations that could permissibly vary in number without resulting in a change in the basic function. Unless defined otherwise, technical and scientific terms used in the following examples have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.
1. Materials and methods
An active 1260, 1260II HPLC (Agilent Inc. of Agilent, USA); shimadzu 20AT (Shimadzu corporation, shimadzu), VORTEX mixer (VORTEX 3), erica (guangzhou) instruments and devices limited; medical centrifuge (CL 5R), hunan instrument laboratory instrument development ltd; pH meter (mettler-toledo instruments (shanghai) ltd); milli-Q ultrapure water system, mercury Mills Ltd.
Methanol, acetonitrile: chromatographically pure (sigma aldrich trade ltd) glacial acetic acid, formic acid, phosphoric acid: analytically pure (Guangzhou chemical Co., ltd.).
Column ECOSILC18-eco100A C column (4.6X 250mm,5 μm) (Guangzhou Chlorella) -MYCOTOX C18 (4.6X 250mm,5 μm) (pickering Co.); ZORBAX Eclipse Plus C18 (4.6X 250mm,5 μm) (Agilent); middle spectrum red RD-C18 (4.6X 150mm,5 μm) (middle spectrum technology)
Standard substance: 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol and 10-shogaol (purity not less than 98%, chengdu Podfield biology Co., ltd.) were synthesized to obtain capsaicin (purity 99.5%, shanghai Anshan Sco laboratory science and technology Co., ltd.).
The 9 batches of ginger juice brown sugar samples were purchased from a large supermarket (see table 1), and all sugar samples were stored in plastic jars, cooled in the shade and protected from light, prior to analysis.
TABLE 1 sources of ginger juice brown sugar samples
2. Experimental methods and results
2.1 chromatographic conditions:
a chromatographic column: ECOSILC18-eco100A C column (4.6X 250mm,5 μm); detection wavelength: 280nm; column temperature: 35 ℃; flow rate: 1.0mL/min; sample introduction amount: 20 mu L of the solution; mobile phase: (a) 1% glacial acetic acid, B acetonitrile, gradient elution: 0 to 15.0min,52.0 percent of A; 15.0-17.0min, 52.0% A → 40.0% A; 17.0-43.0min, 40.0% A → 20.0% A; 43.0-43.01min, 20.0% by weight A → 52.0% by weight A;43.01 to 50.0min,52.0% by weight A.
2.2 preparation of test solutions
Accurately weighing 1.00g of ginger sugar, dissolving with appropriate amount of methanol, vortexing for 1min, performing ultrasound for 30min, and diluting to 10ml with methanol. Shaking and filtering with 0.22 μm microporous membrane to obtain test solution.
2.3 Standard solution preparation
Standard stock solutions: respectively and precisely weighing 10mg of each of 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol, 10-shogaol and synthetic capsaicin standard substances, dissolving with methanol, respectively metering to 100mL to obtain standard stock solutions with the concentration of each standard substance of 100 mu g/mL, storing in a refrigerator at the temperature of-20 ℃, and gradually diluting to the required concentration by using a standard substance solvent when in use.
Serial mixed standard solutions: appropriate amount of the standard stock solution is respectively taken and mixed, the mixture is evenly mixed by a micro vortex mixer, and the mixture is diluted into mixed standard substance solution with the concentration of 2-50 mg/l by methanol for linear range investigation and content measurement.
2.4 negative control solution
Brown sugar without ginger was used as a negative control sample, and the corresponding negative sample solution was prepared as under item "2.2".
2.5 chromatographic condition optimization:
2.5.1 selection of extraction method:
the 6 gingerol fractions were soluble in absolute ethanol and methanol while avoiding interference of too high sucrose in the samples. However, absolute ethanol as a solvent differs from the mobile phase, and the baseline shift is large, so methanol is selected as the extraction solvent. The ginger juice brown sugar contains sucrose, gingerol, pigment, sterol, flavone polyphenol, organic acid, etc. In the present study, it was considered that the recovery rate was not a purification method selected, and therefore a longer gradient elution was required to achieve separation to eliminate interference.
2.5.2 Mobile phase selection
Referring to the 2020 edition of chinese pharmacopoeia and the national standard (see table 2), 1% acetic acid-acetonitrile, 0.1% phosphoric acid-acetonitrile, 0.1% formic acid-acetonitrile, 0.1% phosphoric acid-methanol + acetonitrile (1+1), methanol water, and acetonitrile water were tried, and it was found that separation of 7 components could be achieved under the conditions of acetonitrile + acid water. The peaks of methanol water and acetonitrile water are smeared and overlapped. Under the methanol acid water system, the complete peak-off time is longer than 60min. So 1% acetic acid-acetonitrile in accordance with the standard was selected as the mobile phase.
TABLE 2 study of gingerol in pharmacopoeia and standards
2.5.3 detection wavelength selection:
ultraviolet scanning is carried out on 7-component standard solutions at the wavelength of 190-400 nm (the experimental result is shown in figure 1), strong absorption is found at 230nm and 280nm, but the response of each component at 230nm is general through testing, and the response at 280nm is high, and impurities are relatively less. 280nm was chosen as the detection wavelength.
2.6 methodological investigation
2.6.1 specialization examination
As shown in figure 2, 6 gingerol components in the ginger juice brown sugar are identified through comparison, the peak positions are shown as icons, and each quantitative peak is well separated without obvious interference.
2.6.2 Linearity and Range
Precisely sucking a certain volume of the mixed reference substance stock solution, diluting the mixed reference substance stock solution into standard solutions with different mass concentrations according to a multiple relation, determining by adopting an optimized pretreatment method and chromatographic conditions, and drawing a standard curve by taking the peak area average value (y) of 7 component chromatographic peaks as a vertical coordinate and the corresponding standard solution concentration (x) as a horizontal coordinate; the instrument detection Limit (LOD) was determined with a signal-to-noise ratio S/N = 3. The results are shown in Table 3, the linear relationship of the 7 components in the respective concentration ranges is good, and the correlation coefficients are not less than 0.9998; the detection limit of 6-gingerol is 5.56mg/kg; the detection limit of 8-gingerol is 8.77mg/kg; the detection limit of 10-gingerol is 7.81mg/kg; the detection limit of 6-shogaol is 7.11mg/kg; the detection limit of 8-shogaol is 9.43mg/kg; the detection limit of 10-shogaol is 8.57mg/kg. The method has high sensitivity, and can meet the requirement of detecting gingerol in sugar products.
TABLE 3 Standard Curve, linear Range and detection Limit for the 7 Components
2.6.3 Instrument precision test
The mixed control solution was continuously injected for 8 times under the chromatographic conditions of item 2.1, and the results are shown (table 4): the RSD of the peak areas of the synthesized capsaicin of 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol and 10-shogaol is respectively 1.4%, 0.71%, 2.3%,0.92%,1.8%,0.71% and 0.72%, which indicates that the precision of the instrument is good.
TABLE 4 Instrument precision test results
2.6.4 stability test
Preparing a blank labeled (5 mg/l) part according to the sample processing method under item 2.2, placing under the refrigeration condition (2-4 ℃), taking out respectively at 0,1,3,6, 9, 12 and 24h, performing mass content measurement according to the method under item 2.1, and inspecting the stability of the sample. The results show stability under refrigeration for 24h, see table 5.
TABLE 5 24h stability test results (2-4 ℃ in mg/l away from light)
2.6.5 recovery test
1.00g of brown sugar sample is accurately weighed, 3 levels of reference substances are respectively added, under the optimal extraction condition and chromatographic condition, the recovery rate of gingerol in brown sugar is measured for 3 times in parallel at each concentration, and the result is shown in table 6. As can be seen from Table 6, the average recovery rate of 6-gingerol in brown sugar is 95.8-104.1%, the average recovery rate of 8-gingerol is 89.8-103.0%, the average recovery rate of 6-gingerol is 88.2-102.2%, the recovery rate of 10-gingerol is 94.6-105.3%, and the recovery rate of 8-gingerol is 130.0-134.3%. The recovery rate of 10-shogaol is 80.9-90.8%.
TABLE 6 recovery test results (0 ℃ photophobic unit: ug/g)
2.7 calculation of correction factors
The appropriate amount of each of the series of standard solutions prepared under the term "2.3" was precisely extracted, the peak area of each component was measured by normal injection, and the relative correction factor fk/s of the other 6 components was calculated using the synthesized capsaicin As a reference, and the formula fk/s = fk/fs = (CkAs)/(CsAk), where Ck is the content of the other components, ak is the peak area of the other components, cs is the content of the reference, and As is the peak area of the reference, and the results are shown in table 7.
TABLE 7 relative correction factors for 7 ingredients in brown sugar of ginger juice
2.8 Effect of different instruments, different columns on the correction factor
The results of comparative investigation of the effects of 3 high performance liquid chromatographs and 4 chromatographic columns on the correction factors of 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol and 10-shogaol show that the RSD of the correction factors of 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol and 10-shogaol are respectively 1.64%, 3.53%, 3.89%, 3.83%, 6.05% and 4.51%, which indicates that different instruments and different chromatographic columns have small effects on the correction factors, and the results are shown in Table 8.
TABLE 8 Effect of different instruments, different columns on the correction factor
2.9 durability verification of different flow rates
And taking the mixed reference substance solution for sample injection, and respectively inspecting the relative correction factors of other 6 components when the flow rate is 0.7ml/min, 1.0ml/min and 1.5ml/min and the synthesized capsaicin is used as a reference component according to the method in item 2.1, and inspecting the influence of the flow rate fluctuation on the relative correction factors. The results show that the RSD of each relative correction factor is between 0.66 and 4.58 percent, which shows that the relative correction factor has good durability in the range of 0.7 to 1.5ml/min of fluctuation of the flow rate, and the results are shown in Table 9.
TABLE 9 Effect of different flow rates on relative correction factor
2.10 durability validation of different column temperatures
Taking mixed reference substance solution for injection, and respectively inspecting the relative correction factors of other 6 components at the column temperature of 30 ℃, 35 ℃ and 40 ℃ and taking the synthesized capsaicin as a reference component according to the method in item 2.1, and inspecting the influence of the column temperature on the relative correction factors. The results show that the RSD of each relative correction factor is 0.09% -8.85%, which shows that the relative correction factor has good durability in the range of 5 ℃ of column temperature fluctuation, and the results are shown in Table 10.
TABLE 10 Effect of different column temperatures on relative correction factors
2.11 durability validation at different pH
Taking mixed reference substance solution for injection, respectively inspecting relative correction factors of other 6 components when pH of water phase is 1.5, 2.6 and 3.5 and synthesized capsaicin is used as reference component according to the method in item 2.1, and inspecting influence of pH of mobile phase on the relative correction factors. The results show that each relative correction factor RSD is between 0.25% and 5.13%, which shows that the relative correction factor has good durability in the range of 0.5-1 of the fluctuation change of the pH value of the mobile phase, and the results are shown in Table 11.
TABLE 11 Effect of mobile phase pH on relative correction factor
2.12 locating chromatographic Peak of component to be measured
The invention takes the synthesized capsaicin as an internal standard chromatographic peak, and examines relative retention time values of chromatographic peaks of components to be detected, namely 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol and 10-shogaol under the conditions of 3 high performance liquid chromatographs and 4 chromatographic columns, and the results show that RSD of the relative retention time values of 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol and 10-shogaol are respectively 0.97%, 3.46%, 2.25%, 4.30%, 4.83% and 3.78%, which indicates that different instruments and different chromatographic columns have no significant influence on the relative retention time values, and the results are shown in a table 12.
TABLE 12 Effect of different instruments, different columns on the relative Retention time values
2.13 comparison of the measurement results of the QASM method and the external standard method in the measurement of the content of brown sugar in ginger juice
Taking 9 batches of ginger juice brown sugar samples, preparing 3 sample solutions in parallel for each batch, determining peak areas of all components by adopting a method for sample injection, calculating the contents of 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol and 10-shogaol in the ginger juice brown sugar by adopting a one-test-multiple evaluation method and an external standard method respectively, and obtaining results shown in Table 13. And (3) performing paired sample t test on the content results measured by the external standard method and the QAMS by adopting SPSS25 software, wherein the results show that the measured values of the methods with the P being more than 0.05,2 have no obvious difference.
TABLE 13 results of assay n =3,g/100g
3. Conclusion
As shown in FIG. 2, a relatively large interference peak appears in the first 20min, and the interference may be pigments, polyphenols, flavones, etc. Therefore, the first target peak is generated after the peak output time is prolonged to 20min by a gradient elution mode, and other miscellaneous peaks besides the target peak can be seen in 20min-50min, the target peak and the miscellaneous peaks are difficult to completely separate by pure isocratic elution, and a good correction factor cannot be obtained when the target peak and the miscellaneous peaks are not completely separated, but the baseline separation is finally realized by continuously changing an elution mode to avoid interference.
The research establishes a QAMS quantitative method for 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol and 10-shogaol in ginger juice brown sugar by taking the synthesized capsaicin as a reference substance. The method has good precision, stability, repeatability and durability, and all meet the analysis requirements. By adopting the method, the contents of 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol and 10-shogaol in 9 batches of ginger juice brown sugar are detected, and a t-test analysis result shows that the method is basically consistent with an external standard method measurement result and can be used for measuring the contents of the 6 gingerol components. The research result can provide new ideas and references for quality control and quality evaluation of the ginger juice brown sugar.
The embodiments of the present invention have been described in detail, but the description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention. Any modification, equivalent replacement, and improvement made within the scope of the application of the present invention should be included in the protection scope of the present invention.
Claims (5)
1. A method for determining the content of 6 gingerol components in ginger juice brown sugar by one-test-multiple-evaluation method is a high performance liquid chromatography,
the method is characterized in that ECOSILC18-eco100AC18 is used as a chromatographic column, acetonitrile-1% acetic acid solution is used as a mobile phase, gradient elution is carried out, the flow rate is 1.0ml/min, the column temperature is 35 ℃, and the detection wavelength is 280nm; the method comprises the steps of respectively measuring relative correction factors of six components, namely 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol and 10-shogaol by taking added synthetic capsaicin as a reference substance, and respectively calculating the contents of the six components by using the relative correction factors;
the mobile phase takes 1% acetic acid solution as component A, acetonitrile as component B, and the gradient elution conditions are as follows: 0 to 15.0min,52.0 percent of A; 15.0-17.0min, 52.0% A → 40.0% A; 17.0-43.0min, 40.0% A → 20.0% A; 43.0-43.01min, 20.0% by weight A → 52.0% by weight A; 43.01-50.0min, 52.0% A;
in the method, a sample solution is subjected to the following treatment before sample injection: weighing 1.00g of ginger sugar, dissolving with methanol, performing vortex treatment for 1min, performing ultrasonic treatment for 30min, then performing constant volume treatment to 10ml with methanol, shaking up, and filtering with a 0.22-micron microporous membrane.
2. The method for determining the content of 6 gingerol components in brown sugar containing ginger juice as claimed in claim 1, wherein the ECOSILC18-eco100AC18 chromatographic column has a size of 4.6X 250mm,5 μm.
3. The method for determining the content of 6 gingerol components in ginger juice brown sugar as claimed in claim 1, wherein the sample volume is 20 μ l.
4. The method for determining the content of 6 gingerol components in ginger juice brown sugar as claimed in claim 1, wherein the detection range of 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol and 10-shogaol is 2-50 μ g/ml.
5. The method for determining the content of 6 gingerol components in ginger juice brown sugar as claimed in claim 1, wherein the detection limit of 6-gingerol is 5.56mg/kg; the detection limit of 8-gingerol is 8.77mg/kg; the detection limit of 10-gingerol is 7.81mg/kg; the detection limit of 6-shogaol is 7.11mg/kg; the detection limit of 8-shogaol is 9.43mg/kg; the detection limit of 10-shogaol is 8.57mg/kg.
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| CN108414665A (en) * | 2018-04-23 | 2018-08-17 | 河南科高中标检测技术有限公司 | The assay method of gingerol content in a kind of ginger medicinal material and its preparation |
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| CN108414665A (en) * | 2018-04-23 | 2018-08-17 | 河南科高中标检测技术有限公司 | The assay method of gingerol content in a kind of ginger medicinal material and its preparation |
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