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本文建立了一种以3-甲基-2-苯并噻唑酮腙(MBTH)为氧化剂,以微孔板测定β-葡聚糖酶活力的新方法,该法特别适合于成分复杂的材料中痕量β-葡聚糖酶活力的测定。根据MBTH法测定β-葡聚糖酶解产物还原端基的特点,设计并优化了酶活力测定的关键参数。结果表明,在最适p H 5.0和30℃条件下测定β-葡聚糖酶活力,所得酶的工作浓度范围为0~12.3 m U/m L,饲料中酶的工作浓度范围均为0~2.94 U/g,本法测β-葡聚糖酶和四种饲料样品中酶的平均回收率分别为93.1%、97.5%、104.3%、97.0%、106.4%,β-葡聚糖酶活力检测限为0.37 m U/m L,定量限为1.24 m U/m L,四种饲料酶活力检测限分别为0.10 U/g、0.09 U/g、0.14 U/g、0.09 U/g,定量限分别为0.33 U/g、0.30 U/g、0.46 U/g、0.31 U/g。该法准确、低成本、省时、省力,特别是其灵敏度远高于DNS法,可极大程度上避开饲料中其它成分对测定的干扰。
In this paper, a new method for the determination of β-glucanase activity using 3-methyl-2-benzothiazolone hydrazone (MBTH) as oxidant in a microplate was developed. This method is particularly suitable for complex materials Determination of trace β-glucanase activity. The key parameters of enzyme activity determination were designed and optimized according to the determination of the reduction end groups of β-glucan hydrolyzate by MBTH method. The results showed that the optimal concentration of β-glucanase at pH 5.0 and 30 ℃ was 0-12.3 m U / mL. The concentration of enzyme in feed ranged from 0 ~ 2.94 U / g. The average recoveries of β-glucanase and four feed samples by this method were 93.1%, 97.5%, 104.3%, 97.0% and 106.4%, respectively. The β-glucanase activity The limit of quantification was 0.37 mU / m L and the limit of quantitation was 1.24 mU / m L. The detection limits of the four feed enzymes were 0.10 U / g, 0.09 U / g, 0.14 U / g and 0.09 U / g, respectively. 0.33 U / g, 0.30 U / g, 0.46 U / g, and 0.31 U / g, respectively. The method is accurate, low cost, time-saving, labor-saving, especially its sensitivity is much higher than the DNS method, which can largely avoid the interference of other components in the feed.