【作者】 高岳；

【导师】 罗时石； 葛才林；

【作者基本信息】 扬州大学 ， 生物物理学， 2006， 硕士

【摘要】 辐照品辐照前含菌量的确定,为辐照加工过程中选择适宜的辐照剂量提供依据;提供一种既灵敏、简便,又能快速检测辐照灭菌效果的方法已成为辐照行业目前迫切需要解决的问题。本课题主要开展了两个部分的实验工作。第一部分是:以脱水蔬菜和调味品为材料,进行了样品辐照前含菌量的快速检测方法研究;第二部分是:以脱水蔬菜和调味品为材料,对样品的辐照灭菌效果进行了初步的研究。试验结果如下:1脱水蔬菜和调味品辐照前含菌量的ATP发光快速分析技术研究1.1辐照对菌落总数和大肠杆菌杀菌效果的比较实验脱水蔬菜和调味品中的致病菌主要是大肠杆菌,而从我们的研究结果中可以看出,大肠杆菌的辐照杀死剂量远小于其他普通细菌。因此,只要控制脱水蔬菜和调味品中的细菌总量,就可以保证其达到企业产品质量卫生标准。而ATP生物发光法所研究的对象就是样品中的细菌总数。1.2 ATP的发光强度与样品含菌量的相关性分析ATP的发光强度与ATP浓度之间具有良好的线性关系,相关系数达到0.99。应用细菌ATP生物发光技术测定脱水蔬菜和调味品的辐照前含菌量,从样品的制备到ATP发光强度的测定仅需1~2小时,脱水蔬菜和调味品中细菌ATP的生物发光强度与其含菌量之间呈显著正相关。实验中三种调味品的三条曲线之间的斜率有所不同。分析其原因,除了试验误差外,主要是由于样品溶液带有的颜色不同所导致的光子淬灭程度不同。2脱水蔬菜和调味品辐照灭菌效果的生物发光快速分析技术的初步研究2.1辐照对样品细菌ATP生物发光的影响变化研究样品细菌ATP发光随辐照剂量的增加都比对照有所增加,特别是在5.0kGy时,细菌ATP发光达到最大值,与理论情况:辐照剂量越大,细菌含量越少,ATP发光值也越小不符。这种现象不但在脱水香葱粉中存在,在其他调味品中也同样出现,并不因样品的不同而改变;我们也曾采用三氯醋酸提取法对经辐照后样品中细菌的ATP进行提取,但测试结果变化趋势依旧,说明了ATP生物发光值的升高与提取方法无关;通过对辐照样品的连续跟踪测定,发现这一现象具有较长的时效性,持续达50d左右。压力蒸汽灭菌是目前常用的灭菌方法,我们应用ATP生物发光法进行了121℃压力蒸汽灭菌效果检测的研究,从结果来看,经121℃压力蒸汽灭菌的样品其发光值随着灭菌时间的增加,ATP发光值不断减小;在应用ATP生物发光技术快速测定细菌的药物敏感性时,随着庆大霉素浓度的增加,细菌ATP的发光逐渐变小。都与理论情况相符。综合我们的试验结果和有关报道,与压力蒸汽灭菌和药物灭菌有所不同,γ射线产生的辐射效应已对细菌ATP生物发光的检测产生了影响。2.2辐照后样品细菌ATP发光反应动力学分析发光动力学分析表明,未辐照样品和辐照样品发光曲线变化趋势相同,但辐照后样品的截距Y0大于未辐照样品,而且发光值衰减速率常数k值也高于未辐照的样品,并且差异显著。2.3辐照对样品中细菌ATP的影响研究应用紫外分光光度计对辐照后细菌ATP进行了动态变化的研究,结果显示,随着辐照剂量的增加,细菌ATP含量不断减少;而对ATP标样的研究表明,随着辐照剂量的增加,ATP的发光逐渐降低。综合试验结果可以看出,样品辐照后细菌ATP发光增强不是由于ATP的增加或使ATP处于激发态而引起的,及与ATP本身无关。2.4辐照对细菌ATP生物发光的干扰途径分析对细菌载体和样品细菌洗脱液的研究表明,辐照后ATP发光增强是由电离辐射对细菌的辐射生物学效应产生的代谢产物所造成的,而与细菌的载体无关(本实验中的脱水蔬菜和调味品)。而荧光素—荧光素酶中混有的杂酶,可能是引起干扰的另一个方面的原因。2.5辐照对样品细菌ATP生物发光干扰的特征光谱分析未辐照样品的发光光谱分布呈单峰曲线,其峰值在555nm处,这一谱带为ATP所产生。经6.0kGy辐照处理后样品的ATP发光谱带在555nm处的下降幅度较大,说明辐照后样品中的含菌量已下降, ATP的发光强度将相应下降。但经辐照处理后样品的发光出现了双峰曲线,说明辐照后的样品在ATP正常发光的基础上还存在干扰因素的发光,参照文献我们初步认为脂质过氧化产物很有可能是产生发光干扰的原因,但还有待于进一步研究的确认。更多还原

【Abstract】 In order to keep greenness and quality, reduce expense and prolong store time, we often turn to the method of irradiation. Food irradiation is one method to sterilize, kill pest in food, refrain sprouting , delay ripeness and so on byγray, in order to ensure fresh and quality, reduce loss and prolong storage time. Ensure the primary bacteria number in the sample, then we can choose the fitting radiation dose during irradiation process; In radiation industry, a rapid method for the efficiency of sterilization withγray on samples is a big issue to solve, so we did two aspects of work. First, the rapid-measuring method for primary bacteria number in the sample; second, the rapid bioluminescence assay method for the efficiency of sterilization withγrays.1 Study on the rapid bioluminescence assay method for primary bacteria number in dry vegetable and condiment1.1 Study the efficiency of sterilization comparatively between the total numbers of bacteria and coliform using culture methodColiform is the mostly nosogenesis bacteria in dry vegetable and condiment, the research results show that make the coliform death, the radiation dose is less than other bacteria. If only control the total numbers of bacteria, the sample can reach the sanitation standard.1.2 Relativity analysis between the bioluminescence degree and the bacteria number in the sampleIt is imperious for irradiation industry to apply a simple and rapid method for determining the primary number of bacteria, for it is an important tache during the irradiation process. The results show that there is significant correlation between CFU and ATP bioluminescence, and the primary number of bacteria in dry vegetable and condiment can be rapidly obtained with the bioluminescence method.2 Study on the rapid bioluminescence assay method for the efficiency of sterilization withγray2.1 The changes of bioluminescence after different radiation dose on dry vegetable and condimentWhen the radiation dose increases, the ATP bioluminescence of dry vegetable and condiment also increases. Especially the dose is 5kGy, ATP bioluminescence reach to the most. It is not accord with the theory that the fewer bacteria it is the less ATP bioluminescence will be. Study the factor and mechanism deeply is help to the research.2.2 The changes of bioluminescence after 50 days radiationThe 50 days radiation samples’bioluminescence decreases quickly along with the increase of radiation. When the dose comes to 5kGy, the bioluminescence is very little. It is according with the theory. It shows that the bioluminescence increasing has a long-term effect.2.3 The effect of bioluminescence after three different treatmentsWhen the radiation dose increases, the ATP bioluminescence of dry vegetable and condiment also increases. Especially the dose is 5kGy, ATP bioluminescence reach to the most; After different time sterilization use Pressure cooker, the ATP bioluminescence decrease, when the radiation dose increase, when the time is 20min, the ATP bioluminescence is close to the background;Using different consistence Gentamycin to deal with E.coli, when the consistence increase, the ATP bioluminescence decrease, when the consistence come to 8μg/ml, the ATP bioluminescence is close to the background. The back two treatments is not happen exceptional bioluminescence increasing. So the radiation sample’s ATP bioluminescence increase is a special phenomenon by radiation. It results in the mechanism of radiation. 2.4 The changes of bacteria ATP bioluminescence and ATP standard sample after radiationUse ultraviolet radiation spectrophotometer to mensurate the ATP content in the sample. The results show that the sample’s ATP decrease after radiation, it is accord with the results of Xia shouxuan. ATP standard sample after radiation, the bioluminescence is decrease along with the increasing of radiation dose.2.5 The effect of bioluminescence on radiation samples after using Pressure cooker sterilizationWe mensurate the radiation samples’ATP bioluminescence after sterilization, the results shows that the samples’ATP bioluminescence doesn’t increase when it haven’t bacteria in the samples. We can see in the results that the bioluminescence increasing is not because of the bacteria carrier.2.6 The changes of pure bacteria bioluminescence after radiationThe results show that the pure bacteria’s bioluminescence is increased after radiation, the results is according with the dry vegetable and condiment samples. Here we can say that the bioluminescence increasing is because of the bacteria.更多还原