【摘要】：目前,冠心病日益嚴重地威脅著人類的健康和生命,在歐美等發(fā)達國家冠心病死亡已超過所有癌癥死亡的總和,占總死亡率的27.4%。如果再加上腦中風(fēng)的死亡率,則以動脈粥樣硬化為基礎(chǔ)病變導(dǎo)致的死亡率更高。產(chǎn)生這些疾病的主要因素就是人體內(nèi)甾醇含量的嚴重超標,所以必需限制食用甾醇含量高的食物。本研究利用睪丸酮叢生假單胞桿菌(菌株c.test+act5、菌株c.test+tac、菌株c.test)具有消化甾醇這一特性,以膽固醇為底物篩選其降解甾醇的培養(yǎng)條件,使膽固醇降解酶的活力相對提高,以期為高效降解甾醇類物質(zhì)的工程菌的篩選奠定基礎(chǔ)。主要研究結(jié)果如下: 1、通過對影響膽固醇降解的誘導(dǎo)時間、培養(yǎng)基pH值、誘導(dǎo)溫度、裝液量等因素的研究后,發(fā)現(xiàn)誘導(dǎo)時間、培養(yǎng)基pH值、誘導(dǎo)溫度對三種菌株膽固醇降解酶的活力影響較大,裝液量也有一定的影響。由此篩選出最佳誘導(dǎo)時間14小時、培養(yǎng)基最佳pH7.0、最佳誘導(dǎo)溫度30℃、及最佳裝液量50ml于250ml搖瓶中。根據(jù)上述篩選好的培養(yǎng)條件用平板稀釋法測定三個稀釋度下10~(-7)、10~(-8)、10~(-9)三個菌株的活菌數(shù),同一培養(yǎng)條件下每毫升菌液中菌株c.test+act5中的活菌數(shù)最高達1.275×10~8個,菌株c.test+act中的活菌數(shù)達0.824×10~8個,菌株c.test中的活菌數(shù)最少為0.300×10~8個。 2、本研究同時對睪丸酮叢生假單胞桿菌三個菌株降解膽固醇的發(fā)酵培養(yǎng)基組分進行篩選。結(jié)果表明,不同原料對發(fā)酵液中膽固醇降解酶活力影響相差較大,本研究以有機質(zhì)(酵母膏、蛋白胨、葡萄糖、吐溫)和無機鹽(NaCl、K_2HPO_4、MgSO_4、FeSO_4)分別設(shè)計L_(18)(3~7)和L_9(3~4)正交試驗,通過比較膽固醇降解酶活力的大小獲得有機質(zhì)的最佳配方分別為:5g/L酵母膏、10g/L蛋白胨、5g/L葡萄糖粉、0.5ml/L吐溫。
[Abstract]:At present, coronary heart disease is threatening human health and life day by day. In developed countries, such as Europe and America, the death rate of coronary heart disease has exceeded the sum of all cancer deaths, accounting for 27.444% of the total death rate. If combined with stroke mortality, atherosclerotic disease causes higher mortality. The main cause of these diseases is the severe excess of sterols in the human body, so it is necessary to limit the consumption of foods with high levels of sterols. In this study, we used the characteristic of c.test act5, c.test tac, (c.test) to digest sterol, and used cholesterol as substrate to screen the culture conditions of sterol degradation, so that the activity of cholesterol degrading enzyme was improved. In order to lay a foundation for the screening of engineering bacteria that can efficiently degrade sterols. The main results are as follows: 1. The induction time was found through the study of the factors influencing the degradation of cholesterol, such as the induction time, the pH value of the medium, the induction temperature, the amount of liquid, and so on. The pH value of culture medium and induction temperature had great influence on the activity of cholesterol degrading enzyme of the three strains, and the amount of liquid was also affected to some extent. The optimum induction time was 14 hours, the optimal induction temperature of pH7.0, was 30 鈩，

本文編號：2273516