1 Materials and methods UV-visible spectrophotometer: UV-160A (Shimadzu, Japan); ultrasonic scrubber: MS-5000 (Yamato, Japan); methanol (AR) (Hunan Normal University Reagent Factory); chlorogenic acid standard (Shanghai Imported and packaged). The test material Eucommia ulmoides leaves were collected in Jiangyan Forest Farm in May 1997.
2 Research methods
2.1 wavelength selection according to the nature of chlorogenic acid soluble in water, methanol. Accurately weigh 10.1mg of chlorogenic acid standard, dissolve it in 50% methanol solution and dilute to 10ml and dilute it 10 times, then scan it in the range of 800-200nm, take 50% methanol solution as reference, and have Zui large absorption bee at 325nm. . In addition, the chlorogenic acid standard solution was spotted on a chromatography paper, and after using water as a developing agent, the fluorescent portion (ie, the chlorogenic acid portion) was removed from the ultraviolet lamp, and the remaining portion was ultrasonically extracted with a 50% methanol solution for 15 minutes. Using 50% methanol as a reference, scanning in the range of 800-200 nm, there is a large absorption peak of zui at 345 nm, so 345 nm and 325 nm are selected, and the chlorogenic acid absorbance value is (ΔA=A325-A345). ).
2.2 Preparation of the standard curve accurately weigh the appropriate amount of chlorogenic acid standard, diluted with 50% methanol solution into a standard solution containing chlorogenic acid 5.05, 10.1, 15.15, 20. 2, 30.3, 40.4μg per ml, 50% methanol solution for reference. A345 and A325 values ​​were measured at 325 nm 325 nm, and A (ΔA=A325-A345) was obtained. The concentration C (μg/ml) of the sample was plotted on the abscissa, and the absorbance ΔA was plotted on the ordinate. The A-C standard curve was drawn. The regression equation is obtained as: C = K ΔA + B, where K = 66.247, B = a 0.5808.
2.3 Determination of the sample Weigh accurately 0.5000 g of Eucommia ulmoides leaf powder in a 60 ml test tube, add 40 ml of 50% methanol solution, and then ultrasonically extract for 15 min in an ultrasonic washer and filter. The filter residue was further added with 40 ml of a 50% methanol solution, and ultrasonically extracted for 15 minutes. The above operation was repeated four times (three parallels), and the combined filtrate was adjusted to a volume of 250 ml. The chlorogenic acid content was determined by measuring the A325 and A345 values ​​on a spectrophotometer with 50% methanol as a reference.
3 Results and analysis
3.1 Measurement results
3.2 Stability test The sample was allowed to stand at room temperature for 24 hours and then its absorbance value A was not changed much, indicating that it was stable in 50% methanol solution.
3.3 Recovery rate The standard is formulated into a certain concentration and added to the sample. The content was measured and the recovery was calculated.
4 Conclusions (1) The chlorogenic acid is between 5.05 and 40.4 μg/ml, and its absorbance △A=A325-A345 has a good linear relationship with the concentration.
(2) The content of chlorogenic acid in Eucommia ulmoides leaves was determined by UV-visible spectrophotometry. The stability and reproducibility were good, and the recovery was 104.2%. The method is simple, rapid, accurate and stable, and can be used for the determination of chlorogenic acid content in Eucommia ulmoides leaves.
references
1 Meng Qin et al. Determination of chlorophenol in Eucommia ulmoides by paper chromatography-spectrophotometry. Journal of Yunnan College of Traditional Chinese Medicine, 1994, 17(4): 21-22
2 Zhang Fengyun et al. Comparison of determination methods of chlorogenic acid in Eucommia ulmoides leaves, Journal of Northwest Forestry University, 1996, 11(2): 54-57
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