Different specificity of two types of Pseudomonas lipases for C20 fatty acids with a Δ5 unsaturated double bond and their application for selective concentration of fatty acids
Different specificity of two types of Pseudomonas lipases for C20 fatty acids with a Δ5 unsaturated double bond and their application for selective concentration of fatty acids
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DOI：
10.1263/jbb.101.496
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发表时间：
2006-06-01
影响因子：
2.8
通讯作者：
Shimizu, Sakayu
中科院分区：
文献类型：
--
作者：
Kojima, Yuzo;Sakuradani, Eiji;Shimizu, Sakayu
对两个不同的荧光假单胞菌AK102和HU380产生的两种脂肪酶AK-脂肪酶和HU-脂肪酶分别使用六种含有较高C20脂肪酸的油脂进行了脂肪酸水解特异性的评估，这些脂肪酶包括花生四烯酸(5，8，11，14-二十碳四烯酸，AA，或20：4 omega 6)，双高-伽马-亚麻酸(8，11，14-二十碳三烯酸，DGLA，或20：3 omega 6)，5，8，11，14，17-二十碳五烯酸(EPA或20：5(0))，甲酸(5，8，11，14-二十碳三烯酸，MA，或20：3 omega 9)8，11-二十碳二烯酸(20：2 omega 9)和8，11，14，17-二十碳四烯酸(20：40)。虽然HU-脂肪酶对C20脂肪酸没有表现出任何有或没有Delta 5不饱和键的特异性，但它对4，7，10，13，16，19-二十二碳六烯酸(DHA或22：6 omega3)的反应活性相对较低。相比之下，AK-脂肪酶对具有Delta 5不饱和键的C20脂肪酸的反应较弱。但是，随着反应的进行，AK-脂肪酶的特异性逐渐降低。利用这种脂肪酸的专一性，我们通过脂肪酶催化的水解和尿素包合的方法，从同时含有EPA和DHA的鱼油中浓缩EPA或DHA。以12.2%EPA和6.9%DHA的精制鱼油为原料，用胡脂酶进行水解和尿素包合，分别得到43.1%的EPA和7%的DHA。得到的浓缩总脂肪酸占鳕鱼油中脂肪酸的2.6%。因此，EPA在精炼鳕鱼油的脂肪酸中特别集中，在胡脂肪酶部分水解后尿素包合。而乌贼油经AK-脂肪酶和尿素包合后，其EPA含量从14.2%增加到16.8%，DHA含量从16.3%增加到44.6%。尿素精制总脂肪酸的得率为墨鱼油总脂肪酸的9.4%。因此，DHA尤其集中在AK-脂肪酶部分水解后尿素加成的脂肪酸中。我们的结论是，分别使用HU-脂肪酶和AK-脂肪酶可以方便、廉价地获得EPA和DHA浓缩物。此外，从富含多不饱和脂肪酸的油脂中分离和浓缩C20多不饱和脂肪酸(PUFA)是可能的，无论是否有Delta 5双键，包括这两种脂肪酸。
Two kinds of lipases, AK-lipase and HU-lipase, produced by two different Pseudomonas fluorescens strains, AK102 and HU380, respectively, were evaluated as to fatty acid hydrolysis specificity using six types of oil containing higher amounts of C20 fatty acids such as arachidonic acid (5,8,11,14-eicosatetraenoic acid, AA, or 20:4 omega 6), dihomo-gamma-linolenic acid (8,11,14-eicosatrienoic acid, DGLA, or 20:3 omega 6), 5,8,11,14,17-eicosapentaenoic acid (EPA or 20:5(0), mead acid (5,8,11-eicosatrienoic acid, MA, or 20:3 omega 9), 8,11-eicosadienoic acid (20:2 omega 9) and 8,11,14,17-eicosatetraenoic acid (20:40). Although HU-lipase did not show any specificity for C20 fatty acids with respect to the presence or absence of a Delta 5 unsaturated bond, it exhibited comparatively low reactivity for 4,7,10,13,16,19-docosahexaenoic acid (DHA or 22:6 omega 3). In contrast, AK-lipase was less reactive for C20 fatty acids with a Delta 5 unsaturated bond. However, the specificity of hydrolysis of AK-lipase gradually decreased as the reaction proceeded. Utilizing this fatty acid specificity, we concentrated either EPA or DHA from fish oils containing both EPA and DHA by means of lipase-catalyzed hydrolysis and urea adduction. Hydrolysis and urea adduction of refined cod oil including 12.2% EPA and 6.9% DHA with HU-lipase provided free fatty acids with 43.1% EPA and 7% DHA, respectively. The resulting yield of concentrated total fatty acids comprised 2.6% of the fatty acids from the cod oil. Thus, EPA was particularly concentrated in the fatty acids derived from refined cod oil on partial hydrolysis with HU-lipase followed by urea adduction. On the other hand, hydrolysis of cuttlefish oil with AK-lipase followed by urea adduction increase slightly the EPA composition from 14.2% to 16.8%, and markedly enhanced the composition of DHA from 16.3% to 44.6% in the hydrolyzed fatty acids. The yield of purified total fatty acids by urea concentrate was 9.4% of the fatty acids from the cuttlefish oil. Thus, DHA was particularly concentrated in the fatty acids derived from on partial hydrolysis with AK-lipase followed by urea adduction. We concluded that EPA and DHA concentrates can be easily and inexpensively obtained using HU-lipase and AK-lipase, respectively. Furthermore, it might be possible to separate and concentrate C20 polyunsaturated fatty acids (PUFAs) with or without a Delta 5 double bond from PUFAs rich oils including both fatty acids.