桐粕脂溶性粗毒素测定方法的建立及初步应用
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摘要
桐粕是一种较好的蛋白质资源，在我国蕴藏量非常丰富，充分开发利用这一蛋白质资源具有十分重要的意义。近年来，人们开展了大量的桐粕开发研究工作，并取得了较好的成绩，特别是在脱毒方法、脱毒工艺、毒素提取、毒素成分分离、桐粕营养价值的评定以及桐粕作为蛋白饲料原料在畜牧生产上的应用等方面取得了突破性的进展。但是迄今为止，桐粕毒素成分仍不清楚，桐粕毒素的测定方法未见报道，这就妨碍了桐粕在畜牧生产中的广泛应用。本研究在已有研究的基础上，对桐粕脂溶性粗毒素进行了研究，建立了桐粕脂溶性粗毒素测定方法，应用此方法对不同脱毒处理的脱毒效果和不同产地的桐粕毒素的残留水平进行了检测和比较，并对脱毒桐粕安全性评价进行了初步探讨。
本研究发现，用乙醇溶液对桐粕进行脱毒，然后用乙酸乙酯萃取脱毒溶液所得到的桐粕脂溶性粗毒素具有紫外吸收特征，在270nm处有最大的紫外吸收，并且桐粕脂溶性粗毒素的浓度(Y)与其吸光度(X)之间存在显著的线性相关关系，回归方程为：Y=28.109X+2.4533 R~2=O.9993。在270nm处用紫外分光光度计测得桐粕脂溶性粗毒素溶液的吸光度，根据此回归方程和桐粕脂溶性粗毒素稀释到测定浓度时总体积就可以求得桐粕脱出脂溶性粗毒素量，计算公式为：m=4(28.109A+2.4533)×10~(-2)(g)。用此方法对四个不同脂溶性粗毒素水平的桐粕脱出脂溶性粗毒素量进行测定，与实测值比较，相对偏差小于5％。这证明了此方法可用于桐粕脂溶性粗毒素的检测，而且有较高的准确性。
运用桐粕脂溶性粗毒素测定方法，研究不同处理的脱毒效果，结果表明不同处理脱毒效果不同：(1) 不同脱毒时间，脱毒效果不同。时间越长，脱毒效果越好，每延长48小时，脱毒效果提高约10％；(2) 不同脱毒溶剂浓度的脱毒效果也不一样。浓度越高，脱毒效果越好，75％乙醇溶液和95％乙醇溶液的脱毒效果分别约为50％乙醇溶液的脱毒效果的2倍和3倍；(3) 不同温度，脱毒效果不同。温度越高，脱毒效果越好，但温度升高，脱毒效果增加不显著；(4) 在温度32℃，湿度40％的条件下自然发酵24小时，桐
粕中脂溶性毒素含量降低 10儿
－运用此方法对来自湖南慈利县、永顺县、洪江市三植物油厂的桐粕进行检测，桐粕
毒素残留水平存在差异。来自永顺植物油厂桐粕的毒素残留水平最低，来自洪江植物油
厂的桐粕的毒素残留水平最高。
建立的桐粕脂溶性粗毒素测定方法对脱毒桐粕的安全性评价进行了初步探讨。用小
白鼠饲养试验和急性毒性试验证明脱毒桐粕的安全性，然后应用桐粕脂溶性粗毒素测定
方法对其进行检测。初步研究表明：当100g脱毒桐粕在室温25C下用75％乙醇溶液脱
毒48 ’J＼时，然后用乙酸乙酯革取得到的桐粕脂溶性粗毒素在270urn处测得脱毒溶液吸
光度小于0．329，脱出桐粕脂溶性粗毒素量小于0．4689时，脱毒桐粕就较安全。
桐粕脂溶性粗毒素测定方法的建立，为生产中选择较好的桐粕的脱毒方法、脱毒工
艺和评价脱毒桐粕安全性提供了理论依据。
Tungseed meal is a kind of potential protein,abundant in china. It is of significance to exploit and make full use of it. In recent years,studies on Tungseed meal have been made great progress,especially in the method and technology of extraction and separation,evaluation of nutrition value of Tungseed meal and its application to livestock. Up till now,toxin in Tungseed meal hasn' t been made clear,and no approach to evaluation of toxin in Tungseed meal has been reported,which hindered the use of Tungseed meal in livestock production . Based on previous researches,a method of determination of Ester-soluble crude toxin in Tungseed meal is developed in the present study .The detoxicatibn effects were compared among different treatments with the developed method,the remaining levels of toxin in detoxicated Tungseed meal were determined and evaluation of safety of detoxicated Tungseed meal is investigated.
The results of this study reveals that Ester-soluble crude toxin meal extracted by ethanol solution and ETOAC ( ethyl acetate ) is of ultraviolet absorbance and the spectrogram of Ester-soluble crude toxin has the peak absorption at 270nm,and there is a positive correlation between concentration(Y) and absorbance (X).Y=28. 109X+2. 4533,R2=0. 9993. When the ultraviolet absorbance of Ester-soluble crude toxin solution is determined by UV-spectrophometry,the weight of Ester-solution crude toxion in Tungseed meal can be estimated according to regression formula and total volume of Ester-soluble crude toxin solution . The estimation equation is M=4(28. 109A+2. 4533) X 10-2(g) . A comparison with the true values shows that the measured values of four different Ester-soluble crude toxin level mixed samples by this mean is close,and the bia between true values and the measured values is less than 5%,which suggested that this method may be applicable to the determination
of the Ester-soluble crude toxin and accurate as well.
The application of the developed method to the investigation on the effects of detoxication among different treatments showed that different treatments resulted in different detoxication effects . The longer detoxication lasted,the better the effect was. The effect of detoxication improved by 10% when detoxication prolonged 48 hours. The detoxin solvent concentration also affected the effects of detoxication. The higher the concentration of ethanol solvent,the better the effects of detoxication . The effects of 75% of ethanol solution was about two times better than that of 50% of ethanol and the effects of 95% of ethanol was about three times better than that of 50% of ethanol. The detoxication effects were affected by different temperature. The higher the temperature was,the better the effect of detoxication. .However,the effect of detoxication was increased slowly when the temperature was increased rapidly. Natural fermentation also affected the effects of detoxication . The concentration of Ester-soluble crude toxin in Tungseed meal was decreased by 10% when it was fermented for 24 hours at indoor temperature 32 C humidity 40%.
The content of crude toxin extracted from three kinds of Tungseed meal,which were respectively obtained from the oil plants of Cili County,Yongshun County and Hongjiang County of Hunan Province,was determined in this way. The result showed that there was some difference among those content. The content of crude toxin in Tungseed meal from the oil plant of Yongshun was the lowest,while that of Hongjiang wasthe highest.
The evaluation of the safety of detoxified Tungseed meal was preliminarily investigated with the developed method,the method is also applied to the test of the detoxincated Tungseed meal when the safety of the detoxified Tungseed meal was proved by the feeding experiment and acute toxin experiment of mice,The results suggested that the detoxified Tungseed meal was safe when the absorbance of detoxified solution of ester-soluble crude toxin extracted by ethanol and ethyl acetate at 270nm is less than 0. 329 and the weight
of Ester-soluble crude to
本研究发现，用乙醇溶液对桐粕进行脱毒，然后用乙酸乙酯萃取脱毒溶液所得到的桐粕脂溶性粗毒素具有紫外吸收特征，在270nm处有最大的紫外吸收，并且桐粕脂溶性粗毒素的浓度(Y)与其吸光度(X)之间存在显著的线性相关关系，回归方程为：Y=28.109X+2.4533 R~2=O.9993。在270nm处用紫外分光光度计测得桐粕脂溶性粗毒素溶液的吸光度，根据此回归方程和桐粕脂溶性粗毒素稀释到测定浓度时总体积就可以求得桐粕脱出脂溶性粗毒素量，计算公式为：m=4(28.109A+2.4533)×10~(-2)(g)。用此方法对四个不同脂溶性粗毒素水平的桐粕脱出脂溶性粗毒素量进行测定，与实测值比较，相对偏差小于5％。这证明了此方法可用于桐粕脂溶性粗毒素的检测，而且有较高的准确性。
运用桐粕脂溶性粗毒素测定方法，研究不同处理的脱毒效果，结果表明不同处理脱毒效果不同：(1) 不同脱毒时间，脱毒效果不同。时间越长，脱毒效果越好，每延长48小时，脱毒效果提高约10％；(2) 不同脱毒溶剂浓度的脱毒效果也不一样。浓度越高，脱毒效果越好，75％乙醇溶液和95％乙醇溶液的脱毒效果分别约为50％乙醇溶液的脱毒效果的2倍和3倍；(3) 不同温度，脱毒效果不同。温度越高，脱毒效果越好，但温度升高，脱毒效果增加不显著；(4) 在温度32℃，湿度40％的条件下自然发酵24小时，桐
粕中脂溶性毒素含量降低 10儿
－运用此方法对来自湖南慈利县、永顺县、洪江市三植物油厂的桐粕进行检测，桐粕
毒素残留水平存在差异。来自永顺植物油厂桐粕的毒素残留水平最低，来自洪江植物油
厂的桐粕的毒素残留水平最高。
建立的桐粕脂溶性粗毒素测定方法对脱毒桐粕的安全性评价进行了初步探讨。用小
白鼠饲养试验和急性毒性试验证明脱毒桐粕的安全性，然后应用桐粕脂溶性粗毒素测定
方法对其进行检测。初步研究表明：当100g脱毒桐粕在室温25C下用75％乙醇溶液脱
毒48 ’J＼时，然后用乙酸乙酯革取得到的桐粕脂溶性粗毒素在270urn处测得脱毒溶液吸
光度小于0．329，脱出桐粕脂溶性粗毒素量小于0．4689时，脱毒桐粕就较安全。
桐粕脂溶性粗毒素测定方法的建立，为生产中选择较好的桐粕的脱毒方法、脱毒工
艺和评价脱毒桐粕安全性提供了理论依据。
Tungseed meal is a kind of potential protein,abundant in china. It is of significance to exploit and make full use of it. In recent years,studies on Tungseed meal have been made great progress,especially in the method and technology of extraction and separation,evaluation of nutrition value of Tungseed meal and its application to livestock. Up till now,toxin in Tungseed meal hasn' t been made clear,and no approach to evaluation of toxin in Tungseed meal has been reported,which hindered the use of Tungseed meal in livestock production . Based on previous researches,a method of determination of Ester-soluble crude toxin in Tungseed meal is developed in the present study .The detoxicatibn effects were compared among different treatments with the developed method,the remaining levels of toxin in detoxicated Tungseed meal were determined and evaluation of safety of detoxicated Tungseed meal is investigated.
The results of this study reveals that Ester-soluble crude toxin meal extracted by ethanol solution and ETOAC ( ethyl acetate ) is of ultraviolet absorbance and the spectrogram of Ester-soluble crude toxin has the peak absorption at 270nm,and there is a positive correlation between concentration(Y) and absorbance (X).Y=28. 109X+2. 4533,R2=0. 9993. When the ultraviolet absorbance of Ester-soluble crude toxin solution is determined by UV-spectrophometry,the weight of Ester-solution crude toxion in Tungseed meal can be estimated according to regression formula and total volume of Ester-soluble crude toxin solution . The estimation equation is M=4(28. 109A+2. 4533) X 10-2(g) . A comparison with the true values shows that the measured values of four different Ester-soluble crude toxin level mixed samples by this mean is close,and the bia between true values and the measured values is less than 5%,which suggested that this method may be applicable to the determination
of the Ester-soluble crude toxin and accurate as well.
The application of the developed method to the investigation on the effects of detoxication among different treatments showed that different treatments resulted in different detoxication effects . The longer detoxication lasted,the better the effect was. The effect of detoxication improved by 10% when detoxication prolonged 48 hours. The detoxin solvent concentration also affected the effects of detoxication. The higher the concentration of ethanol solvent,the better the effects of detoxication . The effects of 75% of ethanol solution was about two times better than that of 50% of ethanol and the effects of 95% of ethanol was about three times better than that of 50% of ethanol. The detoxication effects were affected by different temperature. The higher the temperature was,the better the effect of detoxication. .However,the effect of detoxication was increased slowly when the temperature was increased rapidly. Natural fermentation also affected the effects of detoxication . The concentration of Ester-soluble crude toxin in Tungseed meal was decreased by 10% when it was fermented for 24 hours at indoor temperature 32 C humidity 40%.
The content of crude toxin extracted from three kinds of Tungseed meal,which were respectively obtained from the oil plants of Cili County,Yongshun County and Hongjiang County of Hunan Province,was determined in this way. The result showed that there was some difference among those content. The content of crude toxin in Tungseed meal from the oil plant of Yongshun was the lowest,while that of Hongjiang wasthe highest.
The evaluation of the safety of detoxified Tungseed meal was preliminarily investigated with the developed method,the method is also applied to the test of the detoxincated Tungseed meal when the safety of the detoxified Tungseed meal was proved by the feeding experiment and acute toxin experiment of mice,The results suggested that the detoxified Tungseed meal was safe when the absorbance of detoxified solution of ester-soluble crude toxin extracted by ethanol and ethyl acetate at 270nm is less than 0. 329 and the weight
of Ester-soluble crude to
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