| Tagetes erecta,an annual herb of the genus marigold in the composite family,is native to Peru and Mexico in the Americas.It has been introduced and cultivated in China since the1990 s.Because of its rich pigment content,it has become an important production raw material for extracting natural pigment lutein,so it is planted in a large area in China.However,at present,only the flower part of marigold in China is used to extract and prepare single products such as lutein and its derivatives.The level of resource utilization is low,and the yield of lutein obtained by traditional extraction process is low,which unable to meet the growing market demand.How to improve the yield of pigment products and improve the resource utilization of marigold has become an urgent problem for enterprises.In addition,a large amount of fermentation wastewater with strong acidity will be produced in the process of preparing lutein products.This part of the wastewater will be directly discharged,which will not only waste the resources of lactic acid and other effective components,but also pollute the environment.At present,although lactic acid compound acidifier has been put into use in aquaculture,the lactic acid contained in fermented juice has never been utilized.If this part of effective active substances are developed into acidifier products,it will have broad prospects.From the four aspects of marigold fermentation process,drying process,the establishment of fermentation juice characteristic map and the analysis of differential components before and after marigold fermentation,this paper explores the best fermentation and drying process in the process of marigold pretreatment,so as to improve the yield of lutein ester and improve the utilization rate of marigold resources;Establish the characteristic map of fermentation juice,identify the effective components such as lactic acid,and explore the feasibility of developing acidifier from the wastewater produced in the pretreatment of marigold,so as to provide a theoretical basis for the comprehensive utilization of fermentation wastewater;Compare the different components of marigold before and after fermentation by mass spectrometry,and preliminarily explore the changes of components in the fermentation process.The research contents are as follows:(1)Optimization of optimal process parameters for lutein extraction pretreatment of marigold,based on the factory extraction process: Firstly,marigold was fermented and dried to obtain flower residue particles,and then extracted and saponified to finally obtain lutein.The two key processes of fermentation and drying in the lutein extraction process are optimized.Through single factor and orthogonal experiments,the optimum fermentation parameters were as follows: fermentation temperature was 35 ℃,fermentation time was 7days,and the amount of starter was 15 mg/kg;The optimum drying process parameters are as follows: the drying equipment is vacuum drying oven,the drying time is 48 h and the drying temperature is 65 ℃.The experimental results were verified according to this process.Finally,the extraction rate of lutein ester with high content was 14.00 %.(2)The HPLC characteristic map of marigold fermented juice samples was established to identify the effective components such as lactic acid on the premise of ensuring its quality standardization.The final experimental conditions were determined by consulting the literature and experimental comparison.The diamonsil C18 chromatographic column was used for gradient elution with acetonitrile-0.1% phosphoric acid aqueous solution as the mobile phase.The column temperature was 35 ℃,the detection wavelength was 210 nm,the flow rate was 1.0 m L/min,and the injection volume was 10 μL.The HPLC characteristic spectra of 24 batches of marigold fermented juice samples were established,five characteristic peaks were determined,lactic acid was identified as the reference peak,and the relative retention time of each characteristic peak was calculated.The relative retention time of each characteristic peak was ≤ 0.3 %.It is confirmed that the method can be used for the quality control of organic acid components in different batches of marigold fermented juice to ensure that different batches of fermented juice contain organic acid components such as lactic acid and acetic acid that can be used to develop feed acidifier.The feasibility of developing acidifier with lactic acid and other organic acids in fermented juice is preliminarily analyzed by referring to the literature,and the value of recycling and reuse of fermented juice is investigated,so as to provide a prerequisite basis for the subsequent use of marigold fermented juice to develop products and use in aquaculture.(3)The volatile components of marigold before and after fermentation were determined by headspace solid phase microextraction combined with gas chromatography-mass spectrometry(HS-SPME-GC-MS),and then the non-volatile differential chemical components of marigold before and after fermentation were analyzed by high performance liquid chromatography quadrupole time of flight tandem mass spectrometry(HPLC-Q-TOF-MS).The results showed that 73 volatile components and 17 non-volatile components were detected in unfermented flowers;A total of 102 volatile components and32 non-volatile components were detected in the flowers after fermentation.On this basis,by comparing the information of compounds,it was found that there were mainly seven volatile components that changed significantly in fermented flowers,including(R,R)-2,3-butanediol(6.35%-2.87%),tetrahydro-3-furanmethanol(13.39%-5.95%)β-Myrcene(4.04%-1.14%),cyclohexene,1-methyl-4-(1-methylethylenel)-(17.76%-5.75%),2-cyclohexen-1-one,3-methyl-6-(1-methyl enel)-(2.61%-1.45%)and other five substances decreased significantly,while the contents of Phenylethyl Alcohol(2.54%-9.44%)and caryophyllene(4.57%-5.92%)increased.At the same time,compared with the non-volatile components before and after marigold fermentation,it was found that the contents of compounds 1,2,3,4,5,9 and 11 decreased significantly after fermentation,among which compounds 2,4 and 5 decreased most significantly,while the contents of compounds 7,15,20,22 and 32 increased significantly compared with the previous ones.The compounds were further analyzed by modern chromatography according to their mass spectrum cleavage pathway,Compounds 1,2,3,4,5,7,9,11,15,20,22 and 32 were preliminarily identified as sorbitol 、8-hydroxy-3-(2-hydroxyethyl)-1-methoxy-6-methyl-2-pentyl-9,10-dihydroanthracene-9,10-d ione 、6-[8-hydroxy-3-(2-hydroxyethyl)-1-methoxy-6-methyl-9,10-dioxo-9,10-dihydroanthracen-2-yl] caproic acid、8-hydroxy-3-[(2E)-4-(8-hydro xy-1-methoxy-6-methyl-9-oxo-2-pentyl-9H-fluoren-3-yl)-2-en-1-yl]-1-methoxy-7methyl-2-pentyl-9H-fluoren-9-one 、(9Z,11 E,13S,15Z)-13-hydroperoxyoctadeca-9,11,15-trienoic acid、(2,5-dimethoxyphenyl)propanoic acid、Quercetagitrin、Isoquercetin、Queretagetin、2,3-dihydroxy-cyclohexanecarbonyl chloride、3,4-dimethoxybenzoic acid 、 Methyl [3-(4-methoxy-4-oxobutyl)-2-[(4-methoxyphenyl)methyl]-7-methyl-6,8-dioxo-5-phenylisoquinol ine-7-yl] cyclopentanoate. |
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