菜籽饼生物脱毒的微生物筛选复配、脱毒机理与应用效果研究
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摘要
发酵法菜籽饼脱毒方法主要指微生物脱毒方法。微生物数量大，种类多，生长快，易培养，且酶系复杂，几乎可以分解一切天然物质。用微生物制剂进行菜籽饼脱毒，有以下几个特点。第一，酶系复杂，分解硫甙的芥子酶多样，能够水解复杂多变的硫甙。第二，分解中间产物的酶系复杂，能将其彻底分解，并合成微生物自身物质。第三，菜籽饼的其它抗营养因子也能被微生物分解，如植酸、单宁、纤维素、芥子碱等，大大地改变菜籽饼的口感，提高菜籽饼的营养成分的利用率。第四，微生物利用自身代谢作用，可以产生香味物质或香味物质的前体物质，提高菜籽饼脱毒后的香味，所以作者认为微生物脱毒方法是一种很有前途的脱毒方法，应该发展广大。
作者通过一年的室内微生物分离、纯化与脱毒试验和一年的养殖试验，研究了土壤中菜籽饼脱毒微生物的富集和筛选、初选的四十多种微生物纯种的菜籽饼脱毒、属内属间霉菌复配的菜籽饼脱毒、筛选复配FDMRM-1的脱毒效果试验和用FDMRM-1生产的脱毒菜籽饼进行的养殖试验。取得的主要研究结果如下：
1．利用菜籽饼为发酵基质，对A类土、B类土、C类土中菜籽饼脱毒微生物进行富集。在富集过程中A类土的菜籽饼脱毒结果比较理想，GS脱毒率98.4％。总体脱毒效果是A类土＞B类土＞C类土。细菌总数，在A类土中经富集后，增殖4.5倍；在B类土中增殖4.0倍；在C类土中增殖3.4倍。霉菌总数在A类土中经富集后，增殖了10.01倍；在B类土中增殖了11.71倍；在C类土中增殖了11.66倍。放线菌总数在A类土中增殖了1.5倍，在B类土中增殖了2.8倍；在C类土中增殖了3.3倍。酵母菌总数在A类土中增殖了4.0倍；在B类土中增殖7.5倍，在C类土中增殖了9倍。根据微生物数量的变化情况，初步认为GS的脱毒率与脱毒微生物的数量的呈正相关，霉菌的脱毒作用＞细菌的脱毒作用＞放线菌的脱毒作用。并从富集的土壤中分离纯化了四十多种微生物将用于FDMRM的研制。
2．菜籽饼中的主要有毒成分硫代葡萄糖甙有多种结构，在同一样品中多种结构并存。不同品种菜籽饼的硫甙含量与组成不同，同一品种菜籽饼的不同样品的硫甙含量与组成也有差异，这是因为菜籽饼的硫甙含量与组成不仅取决于菜籽的遗传特性，同时也受油菜的栽培条件、菜籽的贮藏条件、油脂的加工条件和菜籽饼贮存条件的影响。也就是对某一菜籽饼品种来说，它的硫甙是复杂多变的。
3．对硫甙有降解作用的微生物可能含有某一类或若干类芥子酶，它只能降解菜籽饼中结构与其相适应的某一种或几种硫甙，结构不相适应的硫甙不能降解。某一微生物的硫甙降解率的最高值(或理论值)，应该是相对应的硫甙在总硫甙中所占的百分比。
不同微生物所分泌的芥子酶系不同，在同一样品的菜籽饼上的GS降解率不同。同一
微生物在不同菜籽饼样品上的GS降解率也不同(因不同样品硫贰组成有差异)。菜籽
饼脱毒微生物复配的实质是一种芥子酶的复配，由于复配后芥子酶系类型增加，则对
应结构的硫贰在总硫贰中所占比例增加，硫贰降解率也就会相应增加。所以属内霉菌
复配，GS降解率有所提高，属间霉菌复配，GS降解率又有大幅度提高。远缘多微生
物复配，达到了较为完美的脱毒效果，GS降解率达到了99%以上，植酸、单片、纤
维素也有很大程度的降解。不同微生物间的芥子酶结构与性能有不同程度的重叠现象。
如在甲微生物中芥子酶的结构类型是abc三类，而在乙微生物中芥子酶的结构类型是
abd，说明有ab的重叠，它们的硫贰降解率的差异主要是在ab百分比基础上。与d的
差异。微生物的亲缘关系越远、芥子酶结构差异越大，芥子酶结构的重叠现象越少，
或越轻，成为远缘微生物复配的理论基础。
在远缘多微生物复配过程中加入了白地霉、酵母菌等四种微生物，它们的GS脱
毒率不高，但可以利用无机态氮合成菌体蛋白，使得菜籽饼脱毒之后蛋白质含量提高
到39.2%，从而提高了脱毒饼的营养价值。
4.以FDMRM一1为脱毒菌剂(远缘多微生物复配)进行了菜籽饼生物脱毒过程
中的微生态研究。证明了在发酵过程中，发酵的环境温度不同，发酵过程中pH值变
化状态不同、发酵品温变化也不同。环境温度在25℃条件下品温变化较理想、GS的
降解率%.8%。
5.提出了菜籽饼脱毒微生物的交替培养分离法，也就是分离霉菌用察氏培养基
一RSM培养基交替培养:分离细菌用牛肉膏蛋白脉培养基一RSM培养基交替培养;
分离酵母菌用麦芽汁培养基一RSM培养基交替培养，可以事半而功倍。
6.用FDMRM一1发酵生产的脱毒饼代替100%豆饼粉加入猪饲料中进行猪的育
肥，日增重5079，料肉比3.30:1，和对照(加入豆饼粉)的饲料饲养效果十分接近(P
>0.01)。月同体品质与内脏重量也基本正常，说明在育肥猪饲料中这种脱毒饼能完全代
替豆饼粉，可以大大降低饲料成本。
7.用FDMRM一1发酵生产的脱毒饼饲养黄粉虫连续三代，无致畸作用。脱毒饼
在黄粉虫饲料中添加量不受限制，可依饲料配比确定。既可降低饲料成本，又可提高
饲料营养，加快幼虫生长速度，增加产卵量，可以明显地提高养虫效益。
Fermentative detoxicating method is microbes detoxicating method Of RSM. Microbes can decompose all the natural material with the characters of great quality, many species, fast growing, easy culture and complicatedly enzymes. There were some characters with detoxicating microbes. First, the enzymes were complicated, complicated and changefull GS could be decomposed. Second, the middle matters could be decomposed by the microbes enzymes, too, and selfish material were composed. Third, Phytin acid, Tannins, Sinapine and Fibre in RSM could be decomposed, nutrition and utilization ratio could be raised. Fourth, some fragrant matters were produced with the microbes self-metabolism, and the taste of detoxicated RSM was changed. The test proved that microbes detoxicating method had a great future.
By the experiments of detoxicating microbes seporation, purification and detoxication of RSM in laboratory and feeding test within two ywers, the detoxicating microbes abundance and purification of RSM in soils, single detoxicating effect of 40 species of microbes, the detoxicating result of moulds in genus, between genera and microbes distant combination, feeding pigs and Tenebrio molitor with FDMRM-1 detoxicated RSM were studied in this paper. The main results are as following:
1. The microbes of RSM detoxicated were abundanced in test, in the A soil, B soil and C soil using RSM as fermentation base, abundancing time 8 weeks, determination every week. B in the A soil increased 4.5 times, in the B soil 4.0 times, in the C soil 3.4 times. M in the A soil increased 10.01 times, in the B soil 11.71 times, in the C soil 11.66 times. A in the A soil increased 1.5 times, in the B soil 2.8 times, in the C soil 3.3 times. Y in the A soil increased 4.0 times, in the B soil 7.5 times, in the C soil 9.0 times. The results of dotoxicating in A soil during the abundance were satisfied. This test proved that action of detoxicating was M >Y > B > A on the basic of microbes change. Microbes of 40 species was sepereted and purified from the abundanced soil with RSM, and that wound be used to study and produce the FDMRM-1.
2. There were plenty of structures in RSM toxic GS which exited in common in one sample. The content and constituent of GS were different in different species of RSM and in different RSM sample of a common species because the content and constituent of GS were decided by not only the inherent characteristics of rape but also their cultivated conditions of rape, stored conditions of rapeseed, processed conditions of plant oil and stored conditions of rapeseed meal. It was that the structures of GS were complicated and changeable to the species of RSM.
3. There was one type thioglucosidase or more in one detoxicating microbe. It could only decomposed the glucosinolate which structure was correspending to the enzyme in RSM. It could no decomposed the GS with different structure. The highest measure of detoxicated GS in one microbe was the percentage correspending structure GS with total GS. The typies of thioglucosidases were different in different microbes so the GS decomposed rates were different in a common RSM sample. The GS decomposed rate of a common microbe was different in fifferent sample of RSM (there were difference in different RSM sample). In fact the combination between microbes was the combination of GS enzymes. Because the typies of thioglucosidase owing to the combination were raised and the
proportion of correspending structure with total GS was increased the detoxicated rate of GS could be raised, the detoxicating rate of GS with M in genus > the detoxicating rate of GS with M between genera. The detoxicating result of many microbes distant combination was sotisfied (99.0%).
The content of protein in detoxiated RSM was increased to 39.2% because of the joining of some microbes which could compose their cell protein from inorganic nitrogen and the nutrition value of detoxicated RSM was raised.
There was interlock appearance among enzymes between microbes. If one microbes GS enzym
作者通过一年的室内微生物分离、纯化与脱毒试验和一年的养殖试验，研究了土壤中菜籽饼脱毒微生物的富集和筛选、初选的四十多种微生物纯种的菜籽饼脱毒、属内属间霉菌复配的菜籽饼脱毒、筛选复配FDMRM-1的脱毒效果试验和用FDMRM-1生产的脱毒菜籽饼进行的养殖试验。取得的主要研究结果如下：
1．利用菜籽饼为发酵基质，对A类土、B类土、C类土中菜籽饼脱毒微生物进行富集。在富集过程中A类土的菜籽饼脱毒结果比较理想，GS脱毒率98.4％。总体脱毒效果是A类土＞B类土＞C类土。细菌总数，在A类土中经富集后，增殖4.5倍；在B类土中增殖4.0倍；在C类土中增殖3.4倍。霉菌总数在A类土中经富集后，增殖了10.01倍；在B类土中增殖了11.71倍；在C类土中增殖了11.66倍。放线菌总数在A类土中增殖了1.5倍，在B类土中增殖了2.8倍；在C类土中增殖了3.3倍。酵母菌总数在A类土中增殖了4.0倍；在B类土中增殖7.5倍，在C类土中增殖了9倍。根据微生物数量的变化情况，初步认为GS的脱毒率与脱毒微生物的数量的呈正相关，霉菌的脱毒作用＞细菌的脱毒作用＞放线菌的脱毒作用。并从富集的土壤中分离纯化了四十多种微生物将用于FDMRM的研制。
2．菜籽饼中的主要有毒成分硫代葡萄糖甙有多种结构，在同一样品中多种结构并存。不同品种菜籽饼的硫甙含量与组成不同，同一品种菜籽饼的不同样品的硫甙含量与组成也有差异，这是因为菜籽饼的硫甙含量与组成不仅取决于菜籽的遗传特性，同时也受油菜的栽培条件、菜籽的贮藏条件、油脂的加工条件和菜籽饼贮存条件的影响。也就是对某一菜籽饼品种来说，它的硫甙是复杂多变的。
3．对硫甙有降解作用的微生物可能含有某一类或若干类芥子酶，它只能降解菜籽饼中结构与其相适应的某一种或几种硫甙，结构不相适应的硫甙不能降解。某一微生物的硫甙降解率的最高值(或理论值)，应该是相对应的硫甙在总硫甙中所占的百分比。
不同微生物所分泌的芥子酶系不同，在同一样品的菜籽饼上的GS降解率不同。同一
微生物在不同菜籽饼样品上的GS降解率也不同(因不同样品硫贰组成有差异)。菜籽
饼脱毒微生物复配的实质是一种芥子酶的复配，由于复配后芥子酶系类型增加，则对
应结构的硫贰在总硫贰中所占比例增加，硫贰降解率也就会相应增加。所以属内霉菌
复配，GS降解率有所提高，属间霉菌复配，GS降解率又有大幅度提高。远缘多微生
物复配，达到了较为完美的脱毒效果，GS降解率达到了99%以上，植酸、单片、纤
维素也有很大程度的降解。不同微生物间的芥子酶结构与性能有不同程度的重叠现象。
如在甲微生物中芥子酶的结构类型是abc三类，而在乙微生物中芥子酶的结构类型是
abd，说明有ab的重叠，它们的硫贰降解率的差异主要是在ab百分比基础上。与d的
差异。微生物的亲缘关系越远、芥子酶结构差异越大，芥子酶结构的重叠现象越少，
或越轻，成为远缘微生物复配的理论基础。
在远缘多微生物复配过程中加入了白地霉、酵母菌等四种微生物，它们的GS脱
毒率不高，但可以利用无机态氮合成菌体蛋白，使得菜籽饼脱毒之后蛋白质含量提高
到39.2%，从而提高了脱毒饼的营养价值。
4.以FDMRM一1为脱毒菌剂(远缘多微生物复配)进行了菜籽饼生物脱毒过程
中的微生态研究。证明了在发酵过程中，发酵的环境温度不同，发酵过程中pH值变
化状态不同、发酵品温变化也不同。环境温度在25℃条件下品温变化较理想、GS的
降解率%.8%。
5.提出了菜籽饼脱毒微生物的交替培养分离法，也就是分离霉菌用察氏培养基
一RSM培养基交替培养:分离细菌用牛肉膏蛋白脉培养基一RSM培养基交替培养;
分离酵母菌用麦芽汁培养基一RSM培养基交替培养，可以事半而功倍。
6.用FDMRM一1发酵生产的脱毒饼代替100%豆饼粉加入猪饲料中进行猪的育
肥，日增重5079，料肉比3.30:1，和对照(加入豆饼粉)的饲料饲养效果十分接近(P
>0.01)。月同体品质与内脏重量也基本正常，说明在育肥猪饲料中这种脱毒饼能完全代
替豆饼粉，可以大大降低饲料成本。
7.用FDMRM一1发酵生产的脱毒饼饲养黄粉虫连续三代，无致畸作用。脱毒饼
在黄粉虫饲料中添加量不受限制，可依饲料配比确定。既可降低饲料成本，又可提高
饲料营养，加快幼虫生长速度，增加产卵量，可以明显地提高养虫效益。
Fermentative detoxicating method is microbes detoxicating method Of RSM. Microbes can decompose all the natural material with the characters of great quality, many species, fast growing, easy culture and complicatedly enzymes. There were some characters with detoxicating microbes. First, the enzymes were complicated, complicated and changefull GS could be decomposed. Second, the middle matters could be decomposed by the microbes enzymes, too, and selfish material were composed. Third, Phytin acid, Tannins, Sinapine and Fibre in RSM could be decomposed, nutrition and utilization ratio could be raised. Fourth, some fragrant matters were produced with the microbes self-metabolism, and the taste of detoxicated RSM was changed. The test proved that microbes detoxicating method had a great future.
By the experiments of detoxicating microbes seporation, purification and detoxication of RSM in laboratory and feeding test within two ywers, the detoxicating microbes abundance and purification of RSM in soils, single detoxicating effect of 40 species of microbes, the detoxicating result of moulds in genus, between genera and microbes distant combination, feeding pigs and Tenebrio molitor with FDMRM-1 detoxicated RSM were studied in this paper. The main results are as following:
1. The microbes of RSM detoxicated were abundanced in test, in the A soil, B soil and C soil using RSM as fermentation base, abundancing time 8 weeks, determination every week. B in the A soil increased 4.5 times, in the B soil 4.0 times, in the C soil 3.4 times. M in the A soil increased 10.01 times, in the B soil 11.71 times, in the C soil 11.66 times. A in the A soil increased 1.5 times, in the B soil 2.8 times, in the C soil 3.3 times. Y in the A soil increased 4.0 times, in the B soil 7.5 times, in the C soil 9.0 times. The results of dotoxicating in A soil during the abundance were satisfied. This test proved that action of detoxicating was M >Y > B > A on the basic of microbes change. Microbes of 40 species was sepereted and purified from the abundanced soil with RSM, and that wound be used to study and produce the FDMRM-1.
2. There were plenty of structures in RSM toxic GS which exited in common in one sample. The content and constituent of GS were different in different species of RSM and in different RSM sample of a common species because the content and constituent of GS were decided by not only the inherent characteristics of rape but also their cultivated conditions of rape, stored conditions of rapeseed, processed conditions of plant oil and stored conditions of rapeseed meal. It was that the structures of GS were complicated and changeable to the species of RSM.
3. There was one type thioglucosidase or more in one detoxicating microbe. It could only decomposed the glucosinolate which structure was correspending to the enzyme in RSM. It could no decomposed the GS with different structure. The highest measure of detoxicated GS in one microbe was the percentage correspending structure GS with total GS. The typies of thioglucosidases were different in different microbes so the GS decomposed rates were different in a common RSM sample. The GS decomposed rate of a common microbe was different in fifferent sample of RSM (there were difference in different RSM sample). In fact the combination between microbes was the combination of GS enzymes. Because the typies of thioglucosidase owing to the combination were raised and the
proportion of correspending structure with total GS was increased the detoxicated rate of GS could be raised, the detoxicating rate of GS with M in genus > the detoxicating rate of GS with M between genera. The detoxicating result of many microbes distant combination was sotisfied (99.0%).
The content of protein in detoxiated RSM was increased to 39.2% because of the joining of some microbes which could compose their cell protein from inorganic nitrogen and the nutrition value of detoxicated RSM was raised.
There was interlock appearance among enzymes between microbes. If one microbes GS enzym
引文
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