| ObjectiveBy using gas chromatography-mass spectrometry(GC-MS)technology to analyze the volatile oil of raw and processed bergamot from Guangdong province,study the change rule of before and after processing bergamot volatile oil components;Study on the quality and feature map of multi number of raw and processed bergamot from Guangdong province,probe the content changes of 5,7-dimethoxy coumarin,hesperidin,5-hydroxymethyl furfural(5-HMF)in different raw and processed bergamot from Guangdong province,establish the characteristics atlas of raw and processed bergamot,to provide experimental basis for its processing mechanism,quality evaluation and control.Through cell experimental technique,study on the effect of growth of gastric mucosal epithelial cells(GES-1)with ethanol extract from different raw and processed samples of bergamot.Preliminary study on the effect of processing process to efficacy on cellular level.Methods1.In accordance with the "Chinese Pharmacopoeia" 2015 edition of the 2204 volatile oil determination method(A method),3 batches of raw and processed bergamot volatile oil were extracted and contents were determined.The volatile oil of different samples were analyzed by GC-MS technology,and the changes of volatile oil composition before and after processing were analyzed.2.High performance liquid chromatography(HPLC)method was developed for the determination of 5,7-dimethoxy coumarin,hesperidin and 5-HMF content in 14 batches of raw bergamot and 12 batches of processed bergamot,analysis of the quality.3.The chromatographic information acquisition different samples by HPLC method,analyze the chromatographic information of 14 batches.of raw and the 12 batches of processed bergamot with similarity evaluation software,comparison of differences before and after processing,establish feature map of raw and processed samples.4.Using four methyl thiazolyl tetrazolium(MTT)method to study the alcohol extract effect on growth of GES-1.Results1.The essential oil content was 0.69ml/100g in deqing raw product(R1),and the essential oil content in the corresponding processed product(P1)was 0.34ml/100g;The essential oil content was 0.57ml/100g in hexiang raw product(R2),and the essential oil content in the corresponding processed product(P2)was 0.26ml/100g;The essential oil content was 0.94ml/100g in gaoyao raw product(R3),and the essential oil content in the corresponding processed product(P3)was 0.79ml/100g.2.R1 sample of 44 component peaks,identified 38 peaks,P1 sample appears 46 component peaks,identified 37 peaks and after processing failed to be detected with nonanal(0.063%),neroli ether(0.116%),L-carveol(0.091%)etc.6,after processing newly produced ingredients with beta elemene(0.114%),viridiflorol(0.067%),alpha cadinol 0.121%etc.5;R2 sample of 51 component peaks,identified 39 peaks,P2 sample appears 75 component peaks,identified 55 peaks and after processing failed to be detected with sabinene(0.143%),trans-2-menthenol(0.089%),neroli ether(0.082%),L-carveol(0.085%),after-processing newly produced ingredients with-alpha cubebene(0.102%)、alpha cadinol(1.039%)、alpha copaene(0.118%)、alpha humulene(0.284%)、cis-beta-farnesene(0.284%)、beta-santalene(0.213%)、β-caryophyllen(0.269%)、methyl linoleate(0.318%)etc.14.R3 sample of 51 component peaks,identified 31 peaks,P3 sample appears 48 component peaks,identified 34 peaks and after processing newly produced ingredients with carvacrol(0.079%),cis-alpha-bisabolene(0.05%)and alpha cadinol(0.088%).R1 sample after processing increased the relative percentage content of the components of 14 species,mainly for delta elemene,gamma-muurolene,gamma elemene,germacrene etc.The relative content of the components to reduce of 17 species,mainly for beta thujene,beta linalool,geraniol,alpha terpineol,alpha citral etc.;R2 sample after processing the relative percentage content of the increase in the composition of 12 kinds of beta bisabolene,beta Cadinene,beta elemene,gamma muurolene etc.There are 22 kinds of relativepercentage reduction,mainly D-limonene and beta pinene,alpha phellandrene,alpha terpinene,beta linalool etc.;R3 sample after processing the relative percentage content of the increase in the composition of 18 kinds of beta thujene,alpha pinene,beta pinene,alpha phellandrene,alpha terpinene,neryl acetate,geranyl acetate and beta bisabolene etc.Reduce the relative percentage content of the components of 13 species,mainly for the p-cymene,beta linalool,nerol,beta cyclocitral,alpha citral etc.3.The contents of 5,7-dimethoxy coumarin in 14 batches of raw products were between 1.5652mg/g and 3.1571mg/g,the mean value is2.1127mg/g.The contents of 5,7-dimethoxy coumarin in 12 batches of processed products were between 1.3501mg/g and 2.6114mg/g,the mean value is 1.9643mg/g.The contents of hesperidin in 14 batches of raw products were between 1.3501mg/g and 2.6114mg/g,the mean value is 1.9643mg/g.The contents of hesperidin in 12 batches of processed products were between 0.2153mg/g and 0.9311mg/g,the mean value is 0.4856mg/g.The contents of 5-HMF in 14 batches of raw products were between 0.0209mg/g and 0.3601mg/g,the mean value is 0.0468mg/g.The contents of 5-HMF in 12 batches of processed products were between 0.0826mg/g and 3.5663mg/g,the mean value is 0.9821mg/g.The mean contents of 5,7-dimethoxy coumarin,hesperidin and 5-HMF in 3 batches of own processing products were 2.1637mg/g,0.3825mg/g and 0.1996mg/g respectively,and the mean contents of 5,7-dimethoxy coumarin and hesperidin were 2.7683mg/g,0.4864mg/g in the 3 batches of corresponding raw products,and 5-HMF in the 3 batches of raw materials were not detected.4.Compared with the control map of 14 batches of raw materials,R7 similarity is 0.817.The similarity of the other 13 batches of the product with the control map were all more than 0.9.Compared with the control map of 12 batches of processed materials,P3 similarity was 0.493,P9 similarity was 0.643,P1 was 0.715,P11 was 0.718,and among the other 8 batches,3 batches of similarity were about 0.8 and 5 batches of similarity is above 0.9.5.When the concentration was 25μg/ml,the growth promoting rate of R1 and P1 alcohol extract on GES-1 cells was the highest,which was 19.55%and 28.16%respectively;When the concentration of administration was 25μg/ml and 50μg/ml,the growth promoting rate of R2 and P2 alcohol extract on GES-1 cells was the highest,which was 17.31%and 26.62%respectively;When the concentration of administration was 25μg/ml and 50μg/ml,the growth promoting rate of R3 and P3 alcohol extract on GES-1 cells was the highest,which was 33.70%and 41.15%respectively.Conclusion1.After processing,the 3 batches of raw products essential oil content were significantly decreased,indicating that the steaming process with the loss of volatile oil.GC-MS analysis found that after the processing of the 3 batches of raw materials,volatile oil in the types and content of a certain change.The contents of D-limonene and γ-terpinene were higher than other components in the volatile oil samples of 3 batches of raw and processed products,both produce aroma.Though the relative contents of D-limonene and y-terpinene before and after processing had a certain change,it was not obvious.It showed that the process of processing may have agreat influence on the total content of volatile oil,but little effect to the relative contents of the main chemical components in volatile oil.After processing,some of the components were not detected,some of the components of the new detection,but the relative contents of these components was relatively low,the components of the new detection were larger molecular weight and higher boiling point.There was a certain change in the total composition of the relative percentage content,but the 3 batches of samples were not completely consistent with the change of the content of the components,the overall change law of R1 sample and R2 sample was that the lower boiling point components contents were reduced,and the high boiling point components contents were elevated,R3 sample were processed with some low boiling point components slightly elevated.The reason for this inconsistency should be further studied.2.The average contents of 5,7-dimethoxy coumarin and hesperidin in raw products were slightly higher than in 12 batches of processed products.By statistical analysis,the difference was not statistically significant(P>0.05).The average contents of 5-HMF in processed products was higher than that in raw materials,by statistical analysis,the difference was statistically significant(P<0.01).The difference of the average contents of 5,7-dimethoxy coumarin and hesperidin in 3 batches of self processed products and the corresponding raw product had no statistical significance(P>0.05),the difference of 5-HMF average contents was statistically significant(P<0.01).The results were identical with the statistical analysis of the 14 raw batches and the 12 processed batches.Explained that processing process had a little effect on contents of 5,7-dimethoxy coumarin and hesperidin,while obvious influence on the contents of 5-HMF.3.The main common peaks were the same of raw and processed feature maps.The similarity between the 14 raw batches HPLC map and the control map were all more than 0.8,showed that the stability of the components of each batch of raw materials was relatively high..There was large difference of the similarity between the 12 processed batches HPLC map and the control map.The causes of the results may be different from the source of different processed products in the market,together with the diversity of processing technology,made the similarity of some samples low.Explained that in the production process of bergamot,scientific and reasonable technological process should be made,to ensure the stability of the quality of processed products.4.The ethanol extracts from the 3 batches of raw and processed products had a promoting effect on the growth of GES-1 cell,and processed products better than raw products,and there was a certain relationship with concentration,prompted that processing process may enhance promoting effect.on GES-1 cell growth of the alcohol extracts. |
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