Abstract




 
   

IJE TRANSACTIONS B: Applications Vol. 32, No. 2 (February 2019) 205-211    Article in Press

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  TEXTURAL AND STRUCTURAL CHARACTERIZATIONS OF MESOPOROUS CHITOSAN BEADS FOR IMMOBILIZATION OF ALPHA-AMYLASE: DIFFUSIVITY AND SUSTAINABILITY OF BIOCATALYST
 
H. Heydarzadeh, S. Gilani, M. Farrokhi, Seyed Mohammad Mahdi Nouri and G. Karimi
 
( Received: October 06, 2018 – Accepted: January 03, 2019 )
 
 

Abstract    In the present study, textural and structural characterizations of chitosan bead for immobilization of alpha amylase were studied in detail by N2 adsorption–desorption, MP, BJH plots and FESEM observations. Pore structure observation revealed chemical activation of chitosan bead by glutaraldehyde can change both the isotherm type of adsorption and pores shape. In consistence with textural analysis the high value of pore volume distribution with range of mesopores region indicates the porosity of activated chitosan bead was uniformly increased. Intra-particle diffusion model indicate 97.6% of amylase was adsorbed inside the mesopores of activated chitosan bead owing to increase in kid (rate constant) and reduce of boundary layer effect on diffusion process. In addition, the stability experiments (pH, storage and thermal stability), enzyme leakage, Ca2+ and salt concentration effects were evaluated for immobilized amylase and compared with its free form. Ca2+ concentration of 1 mM shows an excellent impact on relative activity of amylase in its free and immobilized form. NaCl experiments indicate 84% of amylase was covalently immobilized on activated chitosan beads. Further, the Michaelis–Menten kinetic coefficients, Km (~0.4mg/ml) and, Vmax(~227 U/mg Enzyme), point out strong affinity and high activity of immobilized enzyme.

 

Keywords    Structural characterization; Chitosan bead; Alpha Amylase; Covalent immobilization

 

چکیده    در این تحقیق خواص بافتی و ساختاری دانه های کیتوزان به منظور تثبیت آنزیم آلفا آمیلاز به وسیله ی جذب و واجذب نیتروژن، MP، BJH و مشاهدات FESEM مورد مطالعه قرار گرفت. مشاهدات ساختاری حفرات نشان داد که فعال سازی شیمیایی دانه کیتوزان توسط گلوتارآلدئید می تواند هم نوع ایزوترم جذب و هم شکل حفرات را تغییر دهد. به طور موافق با تحلیل بافت، میزان بالاي توزیع حجمی حفرات در محدوده mesopores نشان می دهد که تخلخل دانه کیتوزان فعال شده به طور یکنواخت افزایش یافته است. مدل نفوذ درون ذره ای نشان می دهد که به دلیل افزایش kid (ثابت سرعت) و کاهش اثر لایه مرزی بر پدیده نفوذ، 6/97٪ آمیلاز در داخل حفرات دانه فعال شده کیتوزان جذب شده است. همچنین آزمایش های پایداری ( pH، ذخیره سازی و پایداری حرارتی)، نشت آنزیم، اثرات غلظت نمک و یون کلسیم برای آمیلاز تثبیت شده بررسی و با آنزیم آزاد مقایسه گردید. غلظت 1 میلی مولار یون کلسیم تاثیر بسیار خوبی بر فعالیت نسبی آمیلاز در شکل آزاد و تثبیت شده نشان داده است. آزمایشات تاثیر نمک کلرید سدیم نشان می دهد که 84٪ از آمیلاز به صورت کوالانسی بر روی دانه های فعال کیتوزان تثبیت شده است. علاوه بر این ضرایب سینتیکی میکائیلیس- منتنKm (~ 0.4mg / ml) و Vmax (~ 227 U /mg Enzyme)، نشان دهنده تمایل قوی و فعالیت بالای آنزیم تثبیت شده است.

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