单、双菌发酵淡豆豉不同发酵期纤溶酶活性
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摘要
目的采用单菌种或双菌种发酵制备淡豆豉,对比分析不同发酵阶段发酵产物的纤溶酶活性。方法以黄豆和黑豆为发酵基料,采用枯草芽胞杆菌(1号菌)、伞枝梨头霉(2号菌)、米曲霉(3号菌)单菌种和双菌种发酵(包括前酵和后酵)。采用琼脂糖—纤维蛋白平板法测定不同发酵阶段发酵产物的纤溶酶活性。结果以黄豆为基料,以枯草芽胞杆菌单菌种37℃发酵7d,再经42℃后酵3d,其纤溶酶活性达到最高值804.61IU/g,显著高于其以黑豆为基料发酵的纤溶酶活性。1号菌单独发酵的淡豆豉纤溶酶活性高于2号和3号菌单独发酵及三者两两组合发酵淡豆豉的纤溶酶活性。结论以纤溶酶活性为考察指标,淡豆豉最佳发酵条件为:以黄豆为基料,以枯草芽胞杆菌单菌进行发酵,通过37℃前酵7d,再经42℃后酵3d其纤溶酶活性最高。
Objective To prepare sojae semen praeparatum(SSP)fermented by single strain or double strains,and comparatively analyze the activity of fibrinolysin in the fermented products in different stages.Methods Soybeans and black beans were used as the substrates for fermentation.The fermentation process(including pre-and post-fermentation)was carried out in single-or double-strain way with Bacillus subtilis(No.1 strain),Absidia corymbifera(No.2 strain)and Aspergillus oryzae(No.3 strain).The activity of fibrinolysin was measured by using agarose-fibrin plate assay.Results With soybeans as the substrate and Bacillus subtilis as the single strain,when fermented at 37℃for 7 days and then at 42℃for 3 days,the activity of fibrinolysin was up to 804.61 IU/g,significantly higher than that with black beans as the substrate.The activity of fibrinolysin in SSP fermented by single No.1 strain was higher than that by single No.2 strain,single No.3 strain and any two of the stains.Conclusion With the activity of fibrinolysin as the standard of evaluation,the optimal condition for SSP fermentation is as follows:Soybeans as the substrate,Bacillus subtilis as the single strain,pre-fermentation at 37℃for 7 days and post-fermentation at 42℃for 3 days.
Objective To prepare sojae semen praeparatum(SSP)fermented by single strain or double strains,and comparatively analyze the activity of fibrinolysin in the fermented products in different stages.Methods Soybeans and black beans were used as the substrates for fermentation.The fermentation process(including pre-and post-fermentation)was carried out in single-or double-strain way with Bacillus subtilis(No.1 strain),Absidia corymbifera(No.2 strain)and Aspergillus oryzae(No.3 strain).The activity of fibrinolysin was measured by using agarose-fibrin plate assay.Results With soybeans as the substrate and Bacillus subtilis as the single strain,when fermented at 37℃for 7 days and then at 42℃for 3 days,the activity of fibrinolysin was up to 804.61 IU/g,significantly higher than that with black beans as the substrate.The activity of fibrinolysin in SSP fermented by single No.1 strain was higher than that by single No.2 strain,single No.3 strain and any two of the stains.Conclusion With the activity of fibrinolysin as the standard of evaluation,the optimal condition for SSP fermentation is as follows:Soybeans as the substrate,Bacillus subtilis as the single strain,pre-fermentation at 37℃for 7 days and post-fermentation at 42℃for 3 days.
引文
[1]Chinese Pharmacopoeia Commission.Pharmacopoeia of the People′s Republic of China(part 1)[M].Beijing:China Science and Technology Press,2015:328.(in Chinese)国家药典委员会.中华人民共和国药典(一部)[M].北京:中国科技出版社,2015:328.
[2]NIU Linying,DU Hongna,LIU Jiao,et al.Comparative analysis of isoflavone contents in semen sojae praeparatum before and after processing[J].Oybean Sci,2008,27(4):672-678.(in Chinese)牛丽颖,杜红娜,刘姣,等.淡豆豉炮制前后异黄酮组分含量的比较[J].大豆科学,2008,27(4):672-678.
[3]WANG Xinguo,GE Xizhen,BAI Xia,et al.Effects of semen sojae preparatum on lipid metabolism in ovariectomized rat[J].J Chin Med Mater,2003,26(9):652-654.(in Chinese)王鑫国,葛喜珍,白霞,等.淡豆豉对去卵巢大鼠脂代谢的影响[J].中药材,2003,26(9):652-654.
[4]GAO Suli,NIU Liyin,CAO Xiulian,et al.Protective effect of ferroented soybean extraction on myocardial ischemia in mice[J].Hebei Med J,2007,29(9):923-924.(in Chinese)高淑丽,牛丽颖,曹秀莲,等.淡豆豉提取物抗心肌缺血作用的研究[J].河北医药,2007,29(9):923-924.
[5]NIU Liyin,LIU Jiao,CUI Lijian,et al.The protective effect of tempeh on vascular endothelial injury in early atherosclerotic rats[J].Pharmacol&Clin Chin Mater Med,2007,23(5):120-122.(in Chinese)牛丽颖,刘娇,崔力剑,等.淡豆豉对早期动脉粥样硬化大鼠血管皮损伤的保护作用[J].中药药理与临床,2007,23(5):120-122.
[6]GE Xizhen,LIN Qiang,HUO Qing.Effects of sojae semen praeparatumi(SSP)on advanced glycosylation endproducts(AGEs)in aortae and serum levels of lipid peroxide in hyperlipemia rats[J].Pharmacol&Clin Chin Mater Med,2006,22(3):101-103.(in Chinese)葛喜珍,林强,霍清.淡豆豉煎剂对高脂血症大鼠主动脉晚期糖化产物的抑制作用[J].中药药理与临床,2006,22(3):101-103.
[7]GU Jianming,PAN Chunyun.Analytical approach of soybean isoflavones and its evaluation[J].J Shanghai Univ,2007,13(6):65-67.(in Chinese)顾建明,潘春云.大豆异黄酮的测定方法及其评价[J].上海大学学报,2007,13(6):65-67.
[8]ZHANG Shaoping,GUO Yong.Molecular biology of Douchi fibrinolytic enzyme[J].Chem Life,2009,29(3):444-446.(in Chinese)张少平,郭勇.豆豉纤溶酶的分子生物学[J].生命的化学,2009,29(3):444-446.
[9]WANG Chang.Mutation breeding of high-yielding fibrinolytic enzyme strains XY-1and study on its enzyme production conditions[D].Wuhan:Huazhong University of Science and Technology,2013.(in Chinese)王昶.豆豉纤溶酶高产菌株XY-1的诱变育种及产酶条件研究[D].武汉:华中科技大学,2013.
[10]LI Jiangwei,RAN Guoxia,CHEN Xinmei.Isolarion and thrombolytic effect of the fibrinolytic enzyme fromSemen Sojae Preparatumin vitro[J].Chin J Biochem Pharma,1999,20(3):148-150.(in Chinese)李江伟,冉国峡,陈新梅.豆豉溶栓酶的分离纯化及其体外溶栓作用[J].中国生化药物杂志,1999,20(3):148-150.
[11]ZHANG Limei.Screening and preservation of Douchi fibrinolytic enzyme producing strain[J].J Anhui Agri,2012,40(10):6187-6188.(in Chinese)张丽梅.豆豉纤溶酶产酶菌的筛选及菌种保藏[J].安徽农业科学,2012,40(10):6187-6188.
[12]LI Hua,LI Duo,SHEN Lirong,et al.Optimization of pure culture fermentation technology of bacterial Douchi[J].J Chin Cereals&Oils Associat,2009,24(2):50-53.(in Chinese)李华,李铎,沈丽荣,等.细菌型豆豉纯种发酵工艺优化[J].中国粮油学报,2009,24(2):50-53.
[13]LIN Xiaohua.Study on the fermentation conditions and volatile components of Aspergillus douchi[D].Nanchang:Nanchang University,2012.(in Chinese)林晓华.曲霉型豆豉的发酵条件及挥发性成分的研究[D].南昌:南昌大学,2012.
[14]LI Hui.Study on fermented soybean fermented soybean and its flavoring substances[D].Dalian:Dalian Polytechnic University,2009.(in Chinese)李慧.混合型豆豉发酵及其风味物质的研究[D].大连:大连工业大学,2009.
[15]SUN Chengxin,MOU Guangqing,SUN Yuan.Studies of production with multi-strain fermention donchi[J].China Condiment,2007(3):43-46.(in Chinese)孙成行,牟光庆,孙园.豆豉混合菌种制曲工艺的研究[J].中国调味品,2007(3):43-46.
[16]WANG Ping,CHEN Liyan,SUN Yinlin,et al.Study on fibrinolysin activity,anticoagulation and thrombolysis effects of Cucumber Seed[J].Jilin J Tradit Chin Med,2017,37(2):180-182.(in Chinese)王萍,陈丽艳,孙银玲,等.黄瓜籽蛋白纤溶活性、抗凝及溶栓作用研究[J].吉林中医药,2017,37(2):180-182.
[17]FAN Xiaodan,GUO Yong,LIU Liu.Identification of fibrinolytic enzyme-producing strain and purification of fibrinolytic enzyme[J].J South China Univ Technol(Natl Sci),2007,35(4):91-94.(in Chinese)范晓丹,郭勇,刘柳.产纤溶酶菌种的鉴定及纤溶酶的分离纯化[J].华南理工大学学报(自然科学版),2007,35(4):91-94.
[18]HU Yiliang,HAO Yimin,LIU Dehai,et al.Conditions of shake flash fermention for producingβ-glucosidase by Absidia corymbifera[J].Henan Sci,2013,31(9):1377-1379.(in Chinese)胡宜亮,郝益民,刘德海,等.伞枝犁头酶产β-葡萄糖苷酶液体摇瓶发酵条件研究[J].河南科学,2013,31(9):1377-1379.
[19]YANG Shengyu,WANG Gang,SHEN Yonghong.Microbial physiology[M].Beijing:Chemical Industry Press,2007:244.(in Chinese)杨生玉,王刚,沈永红.微生物生理学[M].北京:化学工业出版社,2007:244.
[20]LI Gang,LONG Kai,SU Mingsheng,et al.Preliminary study on dynamic change of microbial flora in fermentation process to′Yellow Cladding′of Sojoe Semen Praeparatum[J].Chin JExper Tradit Med Form,2014,20(11):139-142.(in Chinese)李刚,龙凯,苏明声,等.淡豆豉炮制至“黄衣上遍”过程中微生物菌群动态变化的初步研究[J].中国实验方剂学,2014,20(11):139-142.
[21]YANG Dan,WANG Hongli,MA Danning,et al.Effect of different fermentation strains on total isoflavone content and protease activity in semen sojae praeparatum[J].Mod Chin Med,2016,18(7):826-830.(in Chinese)杨丹,王洪礼,马丹宁,等.不同发酵条件对淡豆豉中总异黄酮含量及蛋白酶活力的影响[J].中国现代中药,2016,18(7):826-830.
[2]NIU Linying,DU Hongna,LIU Jiao,et al.Comparative analysis of isoflavone contents in semen sojae praeparatum before and after processing[J].Oybean Sci,2008,27(4):672-678.(in Chinese)牛丽颖,杜红娜,刘姣,等.淡豆豉炮制前后异黄酮组分含量的比较[J].大豆科学,2008,27(4):672-678.
[3]WANG Xinguo,GE Xizhen,BAI Xia,et al.Effects of semen sojae preparatum on lipid metabolism in ovariectomized rat[J].J Chin Med Mater,2003,26(9):652-654.(in Chinese)王鑫国,葛喜珍,白霞,等.淡豆豉对去卵巢大鼠脂代谢的影响[J].中药材,2003,26(9):652-654.
[4]GAO Suli,NIU Liyin,CAO Xiulian,et al.Protective effect of ferroented soybean extraction on myocardial ischemia in mice[J].Hebei Med J,2007,29(9):923-924.(in Chinese)高淑丽,牛丽颖,曹秀莲,等.淡豆豉提取物抗心肌缺血作用的研究[J].河北医药,2007,29(9):923-924.
[5]NIU Liyin,LIU Jiao,CUI Lijian,et al.The protective effect of tempeh on vascular endothelial injury in early atherosclerotic rats[J].Pharmacol&Clin Chin Mater Med,2007,23(5):120-122.(in Chinese)牛丽颖,刘娇,崔力剑,等.淡豆豉对早期动脉粥样硬化大鼠血管皮损伤的保护作用[J].中药药理与临床,2007,23(5):120-122.
[6]GE Xizhen,LIN Qiang,HUO Qing.Effects of sojae semen praeparatumi(SSP)on advanced glycosylation endproducts(AGEs)in aortae and serum levels of lipid peroxide in hyperlipemia rats[J].Pharmacol&Clin Chin Mater Med,2006,22(3):101-103.(in Chinese)葛喜珍,林强,霍清.淡豆豉煎剂对高脂血症大鼠主动脉晚期糖化产物的抑制作用[J].中药药理与临床,2006,22(3):101-103.
[7]GU Jianming,PAN Chunyun.Analytical approach of soybean isoflavones and its evaluation[J].J Shanghai Univ,2007,13(6):65-67.(in Chinese)顾建明,潘春云.大豆异黄酮的测定方法及其评价[J].上海大学学报,2007,13(6):65-67.
[8]ZHANG Shaoping,GUO Yong.Molecular biology of Douchi fibrinolytic enzyme[J].Chem Life,2009,29(3):444-446.(in Chinese)张少平,郭勇.豆豉纤溶酶的分子生物学[J].生命的化学,2009,29(3):444-446.
[9]WANG Chang.Mutation breeding of high-yielding fibrinolytic enzyme strains XY-1and study on its enzyme production conditions[D].Wuhan:Huazhong University of Science and Technology,2013.(in Chinese)王昶.豆豉纤溶酶高产菌株XY-1的诱变育种及产酶条件研究[D].武汉:华中科技大学,2013.
[10]LI Jiangwei,RAN Guoxia,CHEN Xinmei.Isolarion and thrombolytic effect of the fibrinolytic enzyme fromSemen Sojae Preparatumin vitro[J].Chin J Biochem Pharma,1999,20(3):148-150.(in Chinese)李江伟,冉国峡,陈新梅.豆豉溶栓酶的分离纯化及其体外溶栓作用[J].中国生化药物杂志,1999,20(3):148-150.
[11]ZHANG Limei.Screening and preservation of Douchi fibrinolytic enzyme producing strain[J].J Anhui Agri,2012,40(10):6187-6188.(in Chinese)张丽梅.豆豉纤溶酶产酶菌的筛选及菌种保藏[J].安徽农业科学,2012,40(10):6187-6188.
[12]LI Hua,LI Duo,SHEN Lirong,et al.Optimization of pure culture fermentation technology of bacterial Douchi[J].J Chin Cereals&Oils Associat,2009,24(2):50-53.(in Chinese)李华,李铎,沈丽荣,等.细菌型豆豉纯种发酵工艺优化[J].中国粮油学报,2009,24(2):50-53.
[13]LIN Xiaohua.Study on the fermentation conditions and volatile components of Aspergillus douchi[D].Nanchang:Nanchang University,2012.(in Chinese)林晓华.曲霉型豆豉的发酵条件及挥发性成分的研究[D].南昌:南昌大学,2012.
[14]LI Hui.Study on fermented soybean fermented soybean and its flavoring substances[D].Dalian:Dalian Polytechnic University,2009.(in Chinese)李慧.混合型豆豉发酵及其风味物质的研究[D].大连:大连工业大学,2009.
[15]SUN Chengxin,MOU Guangqing,SUN Yuan.Studies of production with multi-strain fermention donchi[J].China Condiment,2007(3):43-46.(in Chinese)孙成行,牟光庆,孙园.豆豉混合菌种制曲工艺的研究[J].中国调味品,2007(3):43-46.
[16]WANG Ping,CHEN Liyan,SUN Yinlin,et al.Study on fibrinolysin activity,anticoagulation and thrombolysis effects of Cucumber Seed[J].Jilin J Tradit Chin Med,2017,37(2):180-182.(in Chinese)王萍,陈丽艳,孙银玲,等.黄瓜籽蛋白纤溶活性、抗凝及溶栓作用研究[J].吉林中医药,2017,37(2):180-182.
[17]FAN Xiaodan,GUO Yong,LIU Liu.Identification of fibrinolytic enzyme-producing strain and purification of fibrinolytic enzyme[J].J South China Univ Technol(Natl Sci),2007,35(4):91-94.(in Chinese)范晓丹,郭勇,刘柳.产纤溶酶菌种的鉴定及纤溶酶的分离纯化[J].华南理工大学学报(自然科学版),2007,35(4):91-94.
[18]HU Yiliang,HAO Yimin,LIU Dehai,et al.Conditions of shake flash fermention for producingβ-glucosidase by Absidia corymbifera[J].Henan Sci,2013,31(9):1377-1379.(in Chinese)胡宜亮,郝益民,刘德海,等.伞枝犁头酶产β-葡萄糖苷酶液体摇瓶发酵条件研究[J].河南科学,2013,31(9):1377-1379.
[19]YANG Shengyu,WANG Gang,SHEN Yonghong.Microbial physiology[M].Beijing:Chemical Industry Press,2007:244.(in Chinese)杨生玉,王刚,沈永红.微生物生理学[M].北京:化学工业出版社,2007:244.
[20]LI Gang,LONG Kai,SU Mingsheng,et al.Preliminary study on dynamic change of microbial flora in fermentation process to′Yellow Cladding′of Sojoe Semen Praeparatum[J].Chin JExper Tradit Med Form,2014,20(11):139-142.(in Chinese)李刚,龙凯,苏明声,等.淡豆豉炮制至“黄衣上遍”过程中微生物菌群动态变化的初步研究[J].中国实验方剂学,2014,20(11):139-142.
[21]YANG Dan,WANG Hongli,MA Danning,et al.Effect of different fermentation strains on total isoflavone content and protease activity in semen sojae praeparatum[J].Mod Chin Med,2016,18(7):826-830.(in Chinese)杨丹,王洪礼,马丹宁,等.不同发酵条件对淡豆豉中总异黄酮含量及蛋白酶活力的影响[J].中国现代中药,2016,18(7):826-830.