【正文】 摘 要 I 摘 要 從土壤中篩選出一株產(chǎn) β半乳糖苷酶的枯草芽孢桿菌，并對菌株和其產(chǎn)生的β半乳糖苷酶進(jìn)行了測定。單菌落的最佳生長時(shí)間是 10h，最佳接種量是 %，最佳培養(yǎng)時(shí)間 24h；該菌株產(chǎn)生的 β半乳糖苷酶的最適 pH 是 ，最適生長溫度為 37℃ ，酶的穩(wěn)定性較高， 4h 后相對酶活仍能維持 85%以上； Mg2+對酶活性的促進(jìn)作用最強(qiáng) ，當(dāng) 濃度為 10 mmol/L 時(shí)，酶活力 可 達(dá) ％ ， Cu2+對酶活性抑制作用最強(qiáng)，少量 Cu2+（ 1mmol/L）就可使酶失活。 本實(shí)驗(yàn)還進(jìn)行了 β半乳糖苷酶的固定化實(shí)驗(yàn)。 以海藻酸鈉 為栽體、戊二醛為交聯(lián) 劑， 對乳糖酶進(jìn)行固定化。研究了海藻酸鈉濃度、氯化鈣濃度、戊二醛濃度、固定化時(shí)間對酶固定化的影響，并對固定化酶酶促反應(yīng)的最適 pH、溫度等進(jìn)行了測定。結(jié)果表明， 4％的海藻酸鈉，在溫度 37℃ 、 pH6~氯化鈣濃度 1％、戊二醛濃度 %下對乳糖酶的固定化率最高。酶促反應(yīng)特性測定結(jié)果表明，固定化后乳糖酶的穩(wěn)定性增強(qiáng)，最適溫度范圍較非固定化乳糖酶大，最適 pH 范圍與非固定化乳糖酶大致相同。但固定化酶珠體機(jī)械強(qiáng)度較差，尚待改進(jìn)。 關(guān)鍵 詞 ： β半乳糖苷酶 酶活性 酶的固定化Absract II Absract A strain isolated from soil in producing βgalactosidase of Bacillus subtilis, and strains and βgalactosidase were measured which produces. A single colony of the best growth time is 10h, the best inoculum was %, the best culture time 24h。 optimum pH βgalactosidase enzyme produced by this strain is , the optimum temperature is 37℃ , enzyme high stability, 4h after the relative activity can be maintained above 85%。 Mg2+ on the activity of promoting the strongest, when the concentration of 10 mmol/L, the enzyme activity of up to %, Cu2+ inhibition of enzyme activity most, a small amount of Cu2+ (1mmol/L) can inactivate the enzyme. The experiments were also βgalactosidase immobilization experiments. The lactase was immobilized on sodium alginate carrier by crosslink with glutaraldehyde. The effects of flutaraldehyde concentration, amount of the enzyme and temperature immobilization were analyzed. The reaction conditions (optimun pH and temperature) of the immobilized lactase were studied. The results show that the high recovery rate of immobilized enzyme could be obtained under the conditions of % glutaraldegyde, 4% sodium alginate and 37℃ temperature. Keyword: βgalactosidase, enzymes activity, Immobilization of enzyme目 錄 III 目 錄 摘 要 .............................................................................................................................. I Absract .............................................................................................................................II 目 錄 ........................................................................................................................... III 第 1 章 緒論 .................................................................................................................... 1 研究背景 ............................................................................................................ 1 課題研究目的和意義 ........................................................................................... 2 國內(nèi)外研究現(xiàn)狀和發(fā)展趨勢 ................................................................................ 2 β半乳糖苷酶的來源及其特性 .............................................................................. 4 細(xì)菌產(chǎn)生的 β半乳糖苷酶 .......................................................................... 4 酵母菌產(chǎn)生的 β半乳糖苷酶 ....................................................................... 4 霉菌產(chǎn)生的 β半乳糖苷酶 .......................................................................... 4 β半乳糖苷酶在食品工業(yè)中的應(yīng)用 ....................................................................... 5 水解牛乳和其它乳制品中的乳糖 ................................................................ 5 生產(chǎn)低聚半乳糖 (Galactooligosaccharide) ..................................................... 6 分析食品中乳糖含量 ................................................................................. 6 β半乳糖苷酶在制藥工業(yè)中的應(yīng)用 .............................................................. 7 β半乳糖苷酶在酶聯(lián)免疫反應(yīng) (EIA)中的應(yīng)用 ............................................... 7 第 2 章 材料和方法 .......................................................................................................... 8 材料 ................................................................................................................... 8 材料與試劑 ............................................................................................... 8 儀器與設(shè)備 ............................................................................................... 8 培養(yǎng)基 ...................................................................................................... 8 產(chǎn) β半乳糖苷酶菌的篩選 .................................................................................... 9 菌株篩選方法 ............................................................................................ 9 分離純化 ................................................................................................... 9 菌種生理生化鑒定 ..................................................................................... 9 β半乳糖苷酶粗酶液制備 ............................................................................ 9 鄰硝基苯酚（ ONP）標(biāo)準(zhǔn)曲線 ................................................................... 9 β半乳糖苷酶酶活 力 測定 .......................................................................... 10 β半乳糖苷酶酶學(xué)性質(zhì) ....................................................................................... 10 菌種生長曲線測定 ................................................................................... 10 培養(yǎng)時(shí)間對菌株產(chǎn)酶能力的影響 .............................................................. 10 接種量對菌株 BGJ222 產(chǎn)酶能力的影響 .................................................... 10 pH 值對 β半乳糖苷酶活力和穩(wěn)定性的影響 ............................................... 10 溫度對 β半乳糖苷酶活力和穩(wěn)定性的影響。 .............................................11 不同金屬離子 及離子濃度對 β半乳糖苷酶的影響。 ...................................11 第 3 章 β半乳糖苷酶 酶學(xué) 性質(zhì) ........................................................................................ 12 高產(chǎn) β半乳糖苷酶細(xì)菌的篩選 ........................................................................... 12 鄰硝基苯酚（ ONP）標(biāo)準(zhǔn)曲線 ........................................................................... 12 菌株 生長曲線測定 ............................................................................................ 13 培養(yǎng)時(shí)