| Pu'er tea is made of Yunnan large leaf sun-light dried green tea by aging or microorganism fermentation,which is a post fermented tea;the former is commonly known as pu'er raw tea,and the latter is pu'er ripe tea.In the whole working process of pu'er tea,the pile-fermentation is the core process and the key of pu'er ripe tea quality characteristic formation,which are the common crossover process of microbial metabolism,enzymatic oxidation and moisture and heat reaction.In this study,we extracted the dominant microorganisms and the isolation,filter and identification were done.We investigated the transfomation of tea contents and polyphennol oxidases by controlling the single-microbe fermentation conditions,hoping to provide some theoretical and practical basis for the filter of fine microbial species and controllability of pu'er tea modern fermentation technology in the fermentation of pu'er tea.The specific results are as follows:1.We isolated 53 microbes from Pu'er fermented tea and tea piles of natural fermentation,and selected 16 microorganisms which had good growth potential and high oxidation ability by the czapek's medium with tea liquid.We conducted morphological and microscopic observations to F6,F7,F3,F5,B5 and B1,and carried out the molecular identification to the first five species.The results were as follows:Aspergillus chevalieri(F6),Aspergillus tubingensis(F7),Penicillium citrinum 1(F3),Penicillium citrinum 2(F5)and Pseudomonas Putida(B5).2.Large leaf sun dried green tea in the autumn of the year 2014 from Yunnan was as fermentation raw material,four fungi F6,F7,F3,F5 and two bacteria B5,B1 were used to conduct single microbe fermentation test,control group was incubate on similar condition but with non-inoculated microbial,periodic sampling analysis was conducted.Compared to non-inoculated microbial treatment,the results showed that four fungi significantly accelerated the transformation of the content of the tea in the microbial fermentation,besides,they promoted the changing of pure and mild character of tea taste,and soup color as well,while two bacteria relatively had less affecting on the fermented tea samples.3.The biochemical components of six microbial organisms were studied in the pile-fermentation in different days.The results showed that the tea polyphenols of the fungi F6,F7,F3,F5 showed a decreasing trend as the fermentation proceeded,and there was a bigger descent range on the 5th to 10th day.The transformation ability of the fungi was F5>F3>F7>F6.The ester catechins decreased significantly when the non-ester catechins increased first and then decreased significantly.The ester catechins of B5 and B1 increased first and then decreased,which increased by 3.68%and decreased by 1.14%to the end of the fermentation.The conversion rates of tea polyphenols in B5,B1 were 9.35%and 8.87%respectively.4.The content of theaflavins in 6 microbial treatments showed a downward trend,and theaflavins of F6 and F7 decreased the most,and the B1 decreased slightly.There were significant differences between F7 and B1 groups compared with the control group;the contents of thearubigins in F6,F7,F3,F5 decreased significantly on the 5th to 10th day as a whole,and the contents of thearubigins were very low in the middle and later stage of fermentation.The contents of thearmbigins in B5 and B1 decreased first and then increased,and the contents of thearmbigins were 5.25%and 5.35%respectively on the 20th day which were higher than the control group.The contents of theabrownine in F6,F7,F3 and F5 increased to 4.3 times,4.5 times,4.8 times and 5.2 times of the initial fermentation(2.98%);and the contents of theabrownine in B5,B1 increase to 3.06%,3.33%from 2.98%of the initial fermentation.5.The contents of flavonoids of fungal treatment samples decreased first and then increased as a whole in the fermentation,and the flavonoids of F6,F7 fermentation samples decreased by 11.88%,3.56%respectively while F3,F5 increased by 91.65%,106.6%.The flavonoids of B5,B1 decreased first and then increased,which increased by 2.94%and 4.19%respectively.The water extract of fungi group decreased first and then rebounded slightly as a whole,and the contents of water extract in F6,F7,F3 and FS reduced by 2.27%,9.16%,14.37%,14.45%to the end of fermentation while the water extract decreased by 2.46%,2.06%in B5 and B1 respectively.The contents of amino acid decreased first and then rebounded slightly,and different strains have their own characteristic.At the end of fermentation,the contents of amino acid in F6,F7,F3,F5 decreased by 22.02%,28.38%,29.18%,26.79%respectively while the amino acide reduced by 17.15%and 13.14%in B5,B1.6.We conducted polyphenol oxidase enzyme activity detection and Native-PAGE electrophoresis to fermented samples of F3,F5,B5 in different days by using the fresh leaves of british red nine and Fuyun No.6 as a control.The results showed that the 3 microbes of F3,F5and B5 secreted extracellular polyphenol oxidases which were different from fresh leaves in the process of pile fermentation.The enzyme activity of bacteria were lower than fungi.The activity of polyphenol oxidase and process change in microbes were related to the oxidative transformation of polyphenols in tea.As the fermentation proceeded,the polyphenol oxidase activity of F3,F5 was very low on the 5th day,and the enzymatic activity of F3 reached the maximum on the 10th day while the FS reached the maximum on the 15th day.The contents of tea polyphenols,total amount of catechins,theaflavins,thearubigins decreased,and ester catechins decreased significantly while simple catechins and GA increased on the 0 to 10th day,and then decreased sharply;besides,the contents of theabrownine decreased significantly.The overall changing trends of F3,F5 were consistent,but they played different roles at different stages in the tea fermentation process.The activity of polyphenol oxidase in B5 increased first and then decreased,and the activity reached the maximum on the 10th,but the whole enzyme activity was low,and B5 had little effect on the contents of polyphenols and total amount of catechins,besides,the theaflavins decreased significantly,thearubigins decreased slightly,and the contents of theabrownine increased slightly which were lower than CK group. |
PDF Full Text Request