Isolation,Characterization And Bioactivity Evaluation Of Maca Sterol And Sterol Ester And Identification Of Maca Through Lipid Soluble Component

Phytosterol is a kind of steroidal compound,whose parent nucleus is dominated by cyclopentane hydrophenanthrene structure.Phytosterol has a variety of bioactivities,such as lowering cholesterol,anti-inflammatory,anti-bacterial and so on.Phytosterol,one of the bioactive substances of Maca,exists in the form of free or esters with fatty acids.It has been reported that different plants have different kinds of fat-soluble components and the fat-soluble components in the same plant from different regions are different.Recently,many researchers have paid attention to study of the fat-soluble components of plants.They have identified different types of plants,distinguished the same plants from different regions and assessed quality of Maca by detection of fatsoluble and volatile components.To further study the type,content and bioactivity of sterol in Maca and distinguish Maca from different regions according to fat-soluble and volatile components,our research is as follows:(1)Using the liquid-solid ratio,ultrasonic time and ultrasonic power as reference,an optimal extract method of sterols in Maca was established with single factor experiment and response surface design experiment.The UV-vis spectrophotometry was used for quantitative analysis of sterols in Maca.The feasibility of this extraction method of sterol was assessed by methodology.The result suggests that this method was feasible.The yield of sterols in Maca can arrive at 0.0535% when sterols in Maca is extracted with 11 m L/g(v/w)liquid-solid ratio at 204 W ultrasonic power for 40 min which is considered as optimal extract condition.(2)The saponification method and silica gel column chromatography were used to purify Maca sterol extract.Firstly,the saponification method was used to remove a large amount of saponification impurities from petroleum ether extract of Maca.Nhexane was used as the extractant to extract unsaponified substance in Maca sterol.The preliminary purified Maca sterol was obtained.Then,silica gel column chromatography was used to purify free sterols and esterified sterols from Maca.The purified Maca sterol from different origins was analysized by GC-MS,infrared absorption spectrometry(FT-IR)and UV-Vis.The results show that β-sitosterol and campesterol were the main free sterols in Maca.The content of β-sitosterol was 11.21-27.74 mg/100 g and that of campesterol was 5.13-10.51 mg/100 g in these samples.(3)The antioxidant capacity of Maca sterol was evaluated by DPPH elimination capacity,ABTS elimination capacity and iron reduction capacity using ascorbic acid as control group.3-(4-5-dimethylthiazol-2-yl)-5(3-carboxymeth-oxyphenyl)-2-(4-sulfopheny)-2H-tetrazolium(MTS)method was used to evaluate the anti-proliferation effect of Maca sterol on lung cancer A549,liver cancer MMC-7721,colon cancer SW480,breast cancer MDA-MB-231 and leukemia HL-60.The bacteriostatic effects of Maca sterol and sterol ester on Escherichia coli,Staphylococcus aureus,Salmonella enterica and Pseudomonas aeruginosa were tested using ceftazidime and penicillin G sodium as control group.The anti-inflammatory activity of purified Maca sterol and Maca sterol esters was studied by setting up drug-free group and L-NMMA-positive drug group as controls.The results showed that Maca sterol had a good inhibitory effect on Staphylococcus aureus.When the concentration was 500 mmol/L,the inhibitory rate of β-sitosterol reached 80.26% and that of campesterol reached 78.56%.Maca sterol ester had a good inhibitory effect on NO.When the concentration reached 400 mmol/L,the inhibitory rate of Maca sterol ester on NO production reached 89.14%.However,neither Maca sterol nor Maca sterol esters showed strong anticancer activity.(4)In this study,ultrasonic-assisted extraction was used to extract the fat-soluble components of Maca from 7 different origins.These components were identified by gas chromatography-mass spectrometry.By comparing the obtained data,the fingerprint of Maca was established.The similarity evaluation(including Angle cosine method)and chemical pattern recognition(principal component analysis,PCA)were applied to evaluate the fat-soluble components of Maca.The result suggests that there is remarkable difference of the relative content of fat-soluble components in Maca from7 origins and PCA can be used to identify these Maca from different origins according to the relative content of fat-soluble components.This method provides reference for the rapid tracing of Maca and support for evaluation of quality of Maca.