Abstract
Aspergillus niger S-1 was proved to be a good hesperidinase producer which have been selected for naringinase utilization. Enzyme of this strain had good characteristics and purified relative high degree with good recovery by ammonium sulfate or aceton treatment. Results obtained were summarized as follows (1) The enzyme was most active at 60$^{\circ}C$, when the reaction was performed in the pH 4.0 for 30min. Optimum pH for enzyme activity was 5.0 and activity was retained 78％ at pH value 3.5. (2) Hesperidinase activity retained 95％ of its full activity after treatment at 60$^{\circ}C$ for 30min at pH value 4.0., 70％ at 70$^{\circ}C$ and 65％ at 80$^{\circ}C$. Most stable pH of this enzyme was showed 5.0 after treatment for 24hr at 4$^{\circ}C$ (3) Only Magnesium ion activated enzyme reaction, while other metallic ions, Cu$\^$＋＋/, Mn$\^$＋＋/, Pb$\^$＋＋/, Mo$\^$＋＋/, Ag$\^$＋＋/, Hg$\^$＋＋/ inhibited. (4) Eleven fold purification with 35％ recovery was obtained in the case of 60％ aceton treatment and 10-fold purification with 5.6％ recovery was showed with 40％ aceton comparing to the crude extract Enzyme. (5) Crude enzyme precipitated with 0.4-0.6 saturated ammonium sulfate contained 13f6 of the original enzyme activity with 48-fold increase in specific activity and enzyme has been purified 25 fold with a yield 19％ by 0.6-5.8 saturation. (6) Hesperidinase formation was noticeably increased by addition of small amount of orange-peel extraction on the wheat bran medium.
산업상 이용에 적합한 분리 선정된 Naringinase 생산 균주인 Asrpergillus niger S-1은 동시에 Hesperidinase를 강력히 생산함이 확인되었으며 이 균주가 생산하는 Hesperidinase의 효소학적 성질을 요약하면 다음과 같다. 1. PH 4.0에서 30분간 반응 시켰을때 최적 온도는 6$0^{\circ}C$ 이었으며 최적 작용 pH는 5.0이었으나 pH 3.0에서는 70% 활성을 보였다. 2. pH 4.0에서 30분간 열처리 하였을 경우 6$0^{\circ}C$로 열처리한 결과 90%, 7$0^{\circ}C$일 경우 70%, 8$0^{\circ}C$일 경우 65%의 활성을 가지는 비교적 내열성 효소이었다. 그리고 pH에 대한 안정성은 4$^{\circ}C$에서 24시간 보존한 결과 pH 5.0일 경우 가장 안정 하였다. 3. $Mg^{＋＋}$ 이온은 1$\times$$10^{-2}$M 이하부터는 활성화하였으나 Cu$^{＋＋}$ Mn$^{＋＋}$ Pb$^{＋＋}$ Mo$^{＋＋}$ Ag$^{＋＋}$ Hg$^{＋＋}$등은 저go 하였다. 4. 효소를 Aceton 처리할 경우 60% Aceton 처리에서 배양 추출물의 경우 보다 11배 전제되었으며 35%가 회수 되었으며 40%의 Aceton 처리일 경우 10배 정제되었으며 5.6%가 회수 되었다. 5. 배양 추출 효소를 유안으로 염석할 경우 0.4.~0.6 분획 침전 시킬 경우 48배 정제되었으며 처음 효소의 13%가 회수되었고 0.6~0.8포화 분획에서도 19%가 회수되었고 25배 정제되었다. 6. Hesperidinase 생성은 약간량의 밀감 과피 추출물의 첨가로 현저히 증가 하였다.