浆水芹菜营养功能成分与亚硝酸盐变化规律及加工机理研究
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摘要
浆水芹菜是西北地区传统发酵蔬菜,以其制作简单、风味独特、营养丰富并具有保健功能而千百年流传。近年也有类似商品面市,但其制作工艺一般通过民间“引子”(菌种)家庭自制自用,缺乏系统技术优化及理论研究,更难以实现传统优质食品的现代化。本研究在对民间传统浆水芹菜制作工艺调查总结基础上,对自然发酵过程中微生物的优势菌群进行了分离及鉴定,初步筛选了四株优良菌株,在对该四个菌株发酵特性研究的同时,对浆水芹菜发酵过程中浆水芹菜营养功能成分和亚硝酸盐含量的变化规律进行了系统研究;在对浆水芹菜加工机理及工艺优化研究的基础上,提出了风味独特、保质期长的浆水芹菜优化工艺及其产品标准。主要研究结果如下:
     1.对民间传统制作工艺调查、分析、总结,得到典型、成功的制作工艺为:芹菜→沸水热烫(预杀菌)→加面条汤、特制面汤或米汤→敞口发酵3~5天→成品。
     2.发现浆水芹菜自然发酵过程中优势菌群变化规律如下:在发酵初期(1~3天),好氧菌,如枯草芽孢杆菌、酵母菌、醋酸菌占优势,随后是明串珠菌;发酵中期(4~7天)以兼性厌氧乳酸杆菌为主;发酵后期(8~9天)一些耐酸的酵母菌数量增多。此阶段若继续发酵,会产生很浓的酒精味或酵母味,影响产品质量;超过10天后,若未进行杀菌保存,耐酸的霉菌会滋生,引起产品腐烂变质。从自然发酵过程中分离出四株优良乳酸菌L1、L2、L3和L4,经初步鉴定L1为明串珠菌属,L2为短乳杆菌属,L3和L4为植物乳杆菌属。
     3.面汤中含有一定量可溶性糖和淀粉等物质,给微生物生长繁殖提供了有利条件。因此,比较加面汤的芹菜与不加面汤的芹菜发酵过程,可以发现:加面汤的芹菜发酵速度快,产酸量大,加面汤的芹菜发酵第5天产酸量是不加面汤的芹菜发酵同期产酸量的近2倍。
     4.研究发现,敞口发酵与密闭发酵优势菌群的变化趋势基本一致。但敞口发酵杂菌多,前期醋酸含量稍高;密闭发酵前期杂菌少,产生的醋酸含量也少,进入乳酸发酵阶段相对缓慢,但维持乳酸发酵时间比敞口发酵长,乳酸含量高,产品质量稳定,货架期长,但口感不如敞口发酵好。
     在20~35℃范围内,温度越高,乳酸发酵速度越快,乳酸菌生长迅速,但生长周期短;成品非经杀菌保存周期短;温度低于20℃或超过35℃不利于乳酸菌的生长繁殖;温度在25~30℃之间为最佳发酵温度。
     5.自然发酵或接种发酵都会产生亚硝酸盐高峰,亚硝酸盐含量与硝酸还原酶活性相关。硝酸还原酶活性高峰与亚硝酸盐高峰在发酵前期出现,随着酸度的增加,硝酸还原酶活性在pH为4.5时下降,亚硝酸盐含量高峰也消失。
     添加面汤、接入陈浆水、适当热烫及加盐,可明显抑制亚硝酸盐的峰值,在20~35℃,发酵温度越高,pH值下降越快,亚硝酸盐高峰出现的越早,峰值越低。
     接种发酵可明显降低硝酸还原酶活性和亚硝酸盐含量,无论是接入单一菌株还是接入混合菌株都能明显地降低硝酸还原酶活性和亚硝酸盐含量,尤以接入四种混合菌株(L1、L2、L3和L4= 1:1:1:1)效果最佳。发酵成熟的浆水芹菜中亚硝酸盐低于FAO/WHO规定的允许值。
     6.面汤稀释比例为1:1、热烫2~3 min、接入3%的陈浆水和发酵温度在25~30℃时,pH下降快,酸度高,VB2、VC和氨基酸含量较高,但总胡萝卜素含量损失较大。VB2在整个发酵过程中逐渐升高,浆水芹菜中VB2含量是新鲜芹菜中的7.1倍;发酵初期是Vc损失的主要阶段,当发酵环境的pH<4时,Vc处于较稳定的状态,Vc在整个发酵过程中损失50~60%,但较其他方法Vc保存较好;氨基酸与新鲜芹菜相比增加6倍左右。
     7.以VB2、VC和氨基酸评价,不同接种方法和接种发酵菌群对产品营养成份影响明显,总趋势为:接入混合乳酸菌菌株>接入单一乳酸菌>接入陈浆水(引子)发酵>自然发酵。接入混合菌株浆水芹菜中与接入陈浆水发酵的浆水芹菜相比,VB2是1.1倍, VC是2.45倍,氨基酸VC是1.6倍;口感也好于自然发酵和接入陈浆水发酵的浆水芹菜。研究还发现,亚硝酸盐含量降低和营养物质含量增高(除总胡萝卜素下降外)与发酵过程中乳酸菌的生长繁殖规律密切相关:随着乳酸菌数量的增加,亚硝酸盐含量逐渐降低,而Vc和VB2及氨基酸含量逐渐增高。说明乳酸菌与这些物质密切相关。通过这些物质的测定,可以确定浆水菜最佳自然食用和安全食用期。
     8.建立了发酵工艺的数学模型并优化出最佳发酵工艺为:接种量5%,面汤稀释比例2:1,沸水热烫3 min,发酵温度26℃。
     9.对发酵过程中芹菜黄酮进行了研究,结果表明:发酵后有利于黄酮提取,从而发现“发酵萃取”方法。利用超声波提取浆水芹菜中的黄酮物质,比用同样方法提取新鲜芹菜中芹菜黄酮的得率高1.4倍;接种发酵浆水芹菜总黄酮的得率比自然发酵浆水芹菜总黄酮的得率高。
     10.通过面汤对浆水芹菜发酵的影响和优势菌群、营养成分变化规律的研究、分析,提出浆水芹菜加工机理主要是靠面汤的营养作用、微生物的发酵作用、蛋白质的分解作用和热烫作用完成的。
     11.获得产品保藏方法的数学模型并得到最佳保藏方法:贮藏温度0.5℃,苯甲酸钠用量0.3‰,微波(800w)处理时间68s。
Jiangshui celery was a kind of traditional fermentation vegetables of northwest area in China. It was handed down from generation to generation for thousands years because its simple technology, special flavour and rich nutrition. It was generally recognized as healthy function food. Homologous products were sold at market in the recent years. But its processing technology depended on exchanging“starter”(strains) each other.However, it was hard to process this kind of food in modern ways due to lack of systematic reseach on its fermentation theory.
     In this experiment, traditional folk processing technology was investigated and summaryed. The predominant microorganism flora were isolated and identified, and the four desire strains (flora) were screened. Their fermentation characteristics were reseach, in the same time, the changing regularity of nutritional, founctional compositions and nitrite were systematically studied during Jiangshui celery fermentation. The optimum technology and production standard of Jiangshui celery was put forward based on research of Jiangshui celery processing mechanism and technology.The main results including:
     1.Through investigation, analysis and summary of traditional folk processing technology, the successful technology of Jiangshui celery was: celery→blanching(beforehand sterilizing)→flour or rice adhesive→open fermentation 3~5 days→products.
     2. The changing regularity of the preponderant population during the natural Jiangshui celery fermentation was: Bacillus subtilis, yeast, and acetic bacteria were dominant in the first stage of fermentation(1~3days),then leuconostoc. Anaerobicbacteria were the main species in the middle stage(4~7days). Acid-fast yeasts multiplied in the last period (8~9days) and produced alcohol and yeast bite if the fermentation were keeping on in the period. Ten days later, if products were not sterilized, acid-fast mold propagated and bring about the corruption of products.
     L1、L2、L3 and L4 strains were isolated and identified from the fermentation Jiangshui celery, and L1 was leuconostoc, L2 was lactobacillus brevis, L3 and L4 were lactobacillus plantarum.
     3. Flour adhesive contains much nutrient such as soluble saccharide and starch etc, which provide good conditions for microorganism growth and propagation. Found: fermentation speed and acid production of celery that added flour adhesive were higher than that of fermentation medium without flour adhesive, and the acid content in flour adhesive was nearly 2.0 times than that of fermentation medium without flour adhesive at the fifth fermentation day.
     4. The research found that the change of preponderant population was almost same in open and close fermentation. The obvious difference was that at early stage, the infectious microorganisms in open fermentation were more and acetic content was higher than that of closing fermentation. Therefore the lactic acid fermentation of close fermentation started slowly, but its lactic acid fermentation can last long time and the lactic acid content was high. It means that the quality of products was stable,and shelf life was long,but the mouth-feeling was inferior to the products produced by opening fermentation method.
     The change of preponderant population was effected by the temperature. Within 20~35℃,the fermentation speed and the lactic bacteria propagation become quicker with the temperature increasing, but the period of propagation was short. The length for preservation of products were short if products were not sterilized.However, the lactic acid bacteria growth was constrained if temperature exceeds 35℃. 25~30℃was desirable for lactic acid fermentation.
     5. The nitrite peak appeared in both spontaneous fermentation and inoculating fermentation, and nitrite content depended on nitrate reductase activity. The peaks of nitrite and nitrate reductase activity appeared in fermentation earlier stage. With acid content increasing, nitrate reductase activity declined gradually at pH≤4.5, meanwhile the nitrite peaks also disappeared gradually.
     Nitrite peak could be controlled obviously by adding flour adhesive, salt, inoculating mellow Jiangshui and proper blanching. Within 20~35℃,with the fermentation temperature enhancing, the pH decilined quickly, the nitrite peak appeared early and its value was low. Both the peak of nitrate reductase activity and nitrite content could be lowered by inoculation fermentation, no matter single strain or mixed strins were used, among these treatments, the ratio of strains L1:L2:L3:L4 = 1:1:1:1 achieved the best result. The content of nitrate in final fermentation Jiangshui celery was lower than the standard issued by FAO/WHO.
     6. The diluent ratio of Flour adhesive was 1:1, blanching 2~3 min, fermentation temperature 25~30℃, the pH decreased quickly and acidity was high, VB2 content and Vc conservation rate were high, but total carotene had more loss. VB2 was enhanced gradually and its content during fermentation of Jiangshui celery was 7.1times than that of fresh celery. Loss of VC was heavy during the first period of fermentation. At pH<4, VC content was stable, and VC lost for 50~60% during whole fermentation. Amino acid during fermentation was about 6 times than that of fresh celery.
     7. The VB2,Vc and amino acid as evaluation,the effect of different inoculating method and inoculating strains on products nutrition was obvious. The total trend was:inoculating mix strains>inoculating pure strains>inoculating mellow Jiangshui>spontaneous fermentation.The VB2 of Jiangshui celery produced by inoculating four LAB strains was 1.1times than inoculating mellow Jiangshui, Vc content was 2.45times, and amino acid was 1.6 times. Furthermore, the mouth-feeling of inoculating fermentation was better than later two methods.
     It was also found that nitrite and nutrition content increasing (besides total carotene decreases) were closely related to the regularity of LAB growth. While LAB increasing, the content of nitrite reduced and content of VC,VB2 and AA increased. It was showed: LAB was closely related with those materials. Through analysis of those materials, edible and safety optimum period of Jiangshui vegetables was determined.
     8. The mathematical model of fermentation technology was building-up by analysis of the regression orthogonal test. The optimum fermentation technology was: inoculate amount 5%, flour adhesive:water 2:1, blanching in boiling water for 3 minutes, fermentation temperature 26℃.
     9.The change of flavonoids was researched during fermentation. The conclusions showed: the flavonoids yield were high after fermentation and found the method of“fermentation extraction”.The flavonoids yield in fermentation Jiangshui celery by the method of ultrasonic extraction was 1.4 times than flavonoids yield in fresh celery by the same method. The flavonoids yield of inoculate fermentation are higher than treatment of spontaneous fermentations.
     10. Through reseach and analysis on the function of flour adhesive,change regularity of predominant strains and nutrition content in Jiangshui celery, the results showed that the processing mechanism of Jiangshui celery depended on the nutrition function of flour adhesive, microorganism fermentation, protein decomposition and blanching.
     11.The mathematical model of storage method on Jiangshui celery was builded-up through orthogonal experiment, and the optimum preservation conditions were: storage temperature 0.5℃, benzoic acid-Na dose 0.3‰,microwave (800w)treatment 68s. Under this treatment, the nutrient contents preservation rate was high at the same time.
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