Effects Of Different Processing Methods On Intrinsic Quality And Characters Of Cistanche
Sep 22, 2022
Objective: To optimize the processing methods of Cistanche Cistanche, and to study the effects of different processing methods on the intrinsic quality and traits of Cistanche Cistanche.
Methods: Different pre-drying treatments (unsliced and not steamed, unsliced and steamed for 20 min, fresh sliced, steamed for 5 min after fresh slices) and drying methods (80, 70, 60, 50 ℃ drying, natural The contents of echinacoside, verbascoside, cistanoside A, isoverbasin, 2′-acetylverbasoside, tuberoside A and polysaccharide in the Cistanche samples that were sun-dried, dried in the shade, and freeze-dried were determined, and the comprehensive score was used. The processing method of Cistanche was optimized by method, the taste of samples was determined by electronic tongue technology, the texture of samples was indirectly evaluated by rehydration rate and hardness index, and the correlation between them and the content of main chemical components was studied.
RESULTS: The comprehensive scoring method was used to select the best processing method of fresh Cistanche, steaming for 20 min and then drying at 60 ℃. The correlation study found that the taste, texture and other traits were correlated with the content of some components to varying degrees. .
Conclusion: In order to effectively ensure the quality of Cistanche , it is necessary to select appropriate processing methods for processing in the origin, and the character information such as taste and texture can reflect the active ingredient content of the medicinal material to a certain extent, which is convenient for rapid identification of medicinal material quality.
Key words Cistanche; origin processing; phenylethanol glycosides（Phg）; polysaccharides; electronic tongue; characters
Free Sample of Cistanche Extract Powder
Cistanche is the dry scaly stem of Cistanche Deserticola Y. C. Ma or Cistanche Tubulosa (Schenk) Wight, which has the functions of invigorating kidney yang, nourishing essence and blood, moistening bowel, and laxative [1].
The processing of Cistanche has been practiced since ancient times. In "Zhenglei Materia Medica", there is a record of "picking on May 5th and drying in the shade". The current processing methods include drying, salting, and cellaring. Due to the large volume of Cistanche, high sugar content and difficulty in drying, the above method is not only time-consuming, laborious, and easily affected by natural conditions, but also with the passage of time and secondary moisturizing, the active ingredients are easily lost, affecting the quality of medicinal materials [2].
In order to ensure the high quality and uniformity of the medicinal materials of Cistanche, it is necessary to optimize the processing methods of its origin, so as to standardize and standardize it. Based on the experience of medicinal farmers in the production area and the research reports of related scholars on the processing methods of Cistanche in the production area in recent years [3-6], this experiment took the fleshy stem of the fresh Cistanche as the research object to investigate the pre-drying treatment and drying methods of Cistanche, using multiple indicators. To evaluate the quality of Cistanche and optimize the best processing method in the production area, it aims to provide a basis for the standardization of processing in the production area of Cistanche, and to provide a new reference method for the comprehensive evaluation and overall control of the medicinal material quality of Cistanche.
1 Instruments and materials
1.1 Instruments
Agilent 1260 high performance liquid chromatograph (Agilent Company, USA); UV6100S UV spectrophotometer (Shanghai Meipuda Instrument Co., Ltd.); ASTREE II electronic tongue (France Alpha MOS Company); FA2004 analytical balance (Shanghai Sunny Hengping) Scientific Instrument Co., Ltd.); DV215CD Analytical Balance (Ohaus Instrument Co., Ltd.); GY-4 Digital Hardness Tester (Yueqing Aidelberg Instrument Co., Ltd.); SB-3200DTN Ultrasonic Cleaner (Ningbo Xinzhi Biotechnology Co., Ltd.).
1.2 Materials
Echinacea (Lot: Y29M10H84490), Verbasin (Lot: Y21A9H59554), Cistanoside A (Lot: P14N9F74960), Isorrascoside (Lot: Y12D8H50507), 2′-Acetyl Verbasin (Lot: P27N6F6623), Tube Anthoside A (batch number: P26N6F6622) reference substances were purchased from Shanghai Yuanye Biotechnology Co., Ltd., mass fraction ≥ 98%; D-anhydrous glucose reference substance (batch number: 110833-201707, China National Institute for Food and Drug Control); acetonitrile It is of chromatographic grade; the rest of the reagents are of analytical grade. The fresh Cistanche for the experiment was collected from the Cistanche cultivation base of Hongkuisha Industrial Development Co., Ltd. of Inner Mongolia Alxa Cistanche Group in October 2019. Succulent stems with scaly leaves. The harvested fresh Cistanche was evenly mixed and randomly divided into groups. Each group of samples was set to have 3 replicates. The sample treatment methods are shown in Table 1.
2 Methods and results
2.1 Determination of phenylethanol glycosides
2.1.1 Chromatographic conditions: Agilent Eclipse XDB-C18 column (250 mm×4.6 mm, 5 μm) was used, the mobile phase was 0.1% formic acid solution (A)-acetonitrile (B), gradient elution (0 ~ 10 min, 9% ~ 15% B; 10 ~ 40 min, 15% ~ 26% B; 40 ~ 45 min, 26% ~ 9% B;
The flow rate was 1.0 mL/min; the detection wavelength was 330 nm; the column temperature was 30 °C; the injection volume was 10 μL, and the chromatogram is shown in Figure 1.
2.1.2 Preparation of reference substance solution: Precisely weigh an appropriate amount of echinacoside, verbascoside, cistanoside A, isoverbasin, 2′-acetylverbasin, and tuberoside A, and place it in 5 mL In a brown volumetric flask, dissolve with 50% methanol and dilute to volume to prepare a reference solution with concentrations of 0.809, 0.932, 0.185, 0.272, 0.582, and 0.068 mg/mL, respectively.
2.1.3 Preparation of the test solution: Take 0.5 g of Cistanche Tubulosa sample powder (passed through a No. 4 sieve), accurately weigh it, place it in a brown volumetric flask, accurately add 25 mL of 50% methanol, seal it tightly, and weigh it. After immersion for 30 min, ultrasonication (150 W, 40 kHz) for 40 min, let it cool and then make up the weight, shake well, let stand, take the supernatant and filter it with a 0.45 μm microporous membrane.
2.2 Determination of polysaccharides
2.2.1 Preparation of reference substance solution: take 10.03 mg of D-anhydroglucose reference substance, accurately weigh, dissolve in water and dilute to 100 mL to obtain a reference substance solution with a concentration of 100.3 μg/mL .
2.2.2 Preparation of the test solution: take 0.5 g of Cistanche Tubulosa sample powder (passed through a No. 4 sieve), accurately weigh it, put it in a stoppered conical flask, add 25 mL of 80% ethanol, and sonicate at 80 °C. (150 W, 40 kHz) for 30 min, filtered while hot, and repeated extraction once. After the filter residue was evaporated to dryness, 25 mL of water was added, and sonicated for 30 min. , Precisely draw 1 mL of solution and dilute it to 10 mL.
2.2.3 Detection conditions: take 2 mL of the test solution of Cistanche, add 1 mL of 6% phenol reagent, shake well, quickly add 5 mL of concentrated sulfuric acid, shake for 5 minutes, heat in a boiling water bath for 15 minutes, cool, and place at 482 °C. Absorbance was measured at nm. Use water instead of the test solution as a blank control.
2.3 Examination of linear relationship Dilute the reference solution according to a certain ratio to prepare a mixed reference solution of a series of concentrations, and measure according to the above method. Take the concentration of the reference solution as the abscissa, and the peak area or absorbance as the ordinate. Linear regression, the results are shown in Table 2.
2.4 Methodological investigation
2.4.1 Precision test:
Accurately draw each reference solution, and measure it continuously for 6 times according to the methods under "2.1.1" and "2.2.3", and calculate to obtain echinacoside, verbascoside, cistanoside A, isaveraside, 2′ The RSDs of the peak areas of -Acetyl Verbasoside and Tuberoside A and the glucose absorbance were 0.76%, 0.53%, 0.81%, 0.25%, 0.44%, 0.62%, 0. . 49%, indicating that the instrument precision is good.
2.4.2 Repeatability test:
Take 6 copies of the same batch of Cistanche for testing powder, accurately weigh, prepare the test solution according to the methods under "2.1.3" "2.2.2", and press "2.1.1" "2.2". 2.3", the calculated RSDs of echinacoside, verbascoside, cistanoside A, isoverbasin, 2'-acetylverbasoside, tuberoside A, and polysaccharide were 0.82%, respectively. , 1.47%, 0.88%, 1.53%, 0.55%, 0.44%, 1.13%, indicating that the method has good repeatability.
2.4.3 Stability test:
Accurately draw the test solution of Cistanche under item "2.1.3", and inject samples at 0, 2, 4, 8, 12, and 24 h according to the chromatographic conditions under item "2.1.1", and calculate the pine cones. The RSDs of the peak areas of chrysanthemum glycoside, verbascoside, cistanoside A, isoverascoside, 2′-acetylverbasoside, and tuberoside A were 0. 47%, 1. 12%, 0. 42%, 0. 65, respectively. %, 0.57%, 0.47%, indicating that the test solution has good stability within 24 h; accurately absorb the test solution of Cistanche under "2.2.2", press "2.2.3" The detection conditions under the item were measured at 0, 15, 30, 60, 120, and 240 min respectively, and the RSD of the absorbance of the polysaccharide was calculated to be 1.24%, indicating that the test solution was stable within 240 min.
2.4.4 Sample recovery test:
Take 6 parts of Cistanche for testing powder with known component content, each part is 0.25 g, accurately weighed, add
Add the reference solution with the content of each component in the sample, prepare the test solution according to the methods under "2.1.3" "2.2.2", and according to "2.1.1" "2.2. Measured by the method under the item 3”, and calculated to obtain Echinacea
24%, 102. 69%, 101. 13%, 101. 13%, 101. 13%, 101. 13%, 101. 102. 38%, 101. 27%, 99. 94%, 99. 23%, RSD were 0. 98%, 2. 18%, 0. 86%, 1. 26%, 1. 13%, 0. 94%, 2. 34%, indicating that the two methods are accurate and reliable.
2.5 Effects of different processing methods on the content of active components in Cistanche The sum of the contents of echinacoside and verbascoside in the Cistanche samples was higher than 0.30%, which was in line with the 2020 edition of the Chinese Pharmacopoeia [1].
The study found that the total amount of the index components in the whole dried medicinal materials was higher than that of the fresh slices, and the total amount of the index components in the steamed sample RZ3 before drying was the highest. %, which is more than 14 times the level specified in the Pharmacopoeia. It can be seen that the content of active ingredients after steaming and processing of the medicinal materials in the origin is higher. By comparing the effects of sun-drying, shade-drying, blast drying and freeze-drying on the content of active ingredients in unsliced and steamed Cistanche Cistanche, it was found that the content of echinacoside in sample RZ2 was the highest; The contents of '-acetylverbasin and tuberoside A were the highest, the content of Cistanche A in the sample RZ6 was the highest, and the content of polysaccharide in the sample RZ5 was the highest, indicating that different drying methods had different effects on the content of active components in Cistanche Cistanche. It is necessary to use multiple indicators to comprehensively evaluate its quality.
The AHP method was used to determine the weight coefficient of the quality evaluation index of Cistanche, and the weight coefficients of the seven indicators of echinacoside, verbascoside, cistanoside A, isoverbasin glycoside, 2′-acetylverbasic glycoside, piploside A and polysaccharide were calculated respectively. 0.2668, 0.2668, 0.1626, 0.0958, 0.0958, 0.0561, 0.0561.
According to the importance of the measured components in Cistanche, the 7 indicators were divided into 4 levels, and the priority order of each indicator was determined: echinacoside=verbasicose glycosides>cistanoside A>isoverbasin glycosides=2′-acetyl group Verbasin> tuberosin A=polysaccharide, thus constructing the judgment priority moment for pairwise comparison. The random consistency ratio CR is an index to measure whether the obtained weight coefficient is reasonable. In this experiment, CR= 0. 0066 < 0. 10, indicating that the judgment priority matrix for pairwise comparison of indicators conforms to the consistency test.
The coefficient is reasonable and effective, and can be used for comprehensive scoring of processing methods in the optimal production area of Cistanche [7].
In order to make the data uniform, the evaluation indicators need to be normalized, the formula is yi=xi-xmin*xmax-xmin, where yi is the normalized value of the indicator, xi is the actual value of the indicator, and xmax and xmin are the indicators respectively. The maximum and minimum values of the group, and the weighted comprehensive score is calculated according to the formula K = y1×l1+y2×l2+…y6×l6+y7×l7, where K is the comprehensive score value, y1, y2…y6, y7 respectively is the result of the normalization of the contents of echinacoside, verbascoside, cistanoside A, isoverascoside, 2′-acetylverbasic glycoside, tuberoside A, and polysaccharide. Weights. The comprehensive scoring results showed that the overall quality of the sample RZ3 was the best, that is, the whole fresh Cistanche was steamed for 20 min and then dried at 60℃ as the best processing method in the origin. The results are shown in Table 3.
