مقایسه عملکرد باکتری Methylorubrum sp. در حذف ترکیبات نفت خام به‌صورت آزاد و تثبیت‌شده: رویکردی بر پایه فعالیت آنزیم‌های کلیدی
محورهای موضوعی :میترا پارسا 1 , سنبل ناظری 2 *
1 - دانشجوی دکتری، گروه تولیدات و ژنتیک گیاهی، دانشکده کشاورزی، دانشگاه بوعلی سینا، همدان، ایران
2 - دانشیار، گروه تولیدات و ژنتیک گیاهی، دانشکده کشاورزی، دانشگاه بوعلی سینا، همدان، ایران
کلید واژه: آلکان‌مونو‌اکسیژناز, باکتری Methylorubrum sp., تثبیت سلولی, سیتوکروم P450 و لیپاز,
چکیده مقاله :
نشت نفت خام و ماندگاری ترکیبات هیدروکربنی در محیط‌زیست، تهدیدی جدی برای سلامت انسان و اکوسیستم‌ها محسوب می‌شود. این ترکیبات به دلیل سمیت، پایداری بالا و تجمع در منابع آب و خاک، موجب اختلال در زنجیره‌های غذایی و تشدید بحران‌های زیست‌محیطی می‌شوند. زیست‌پالایی با بهره‌گیری از میکروارگانیسم‌ها، راهکاری مؤثر، اقتصادی و سازگار با محیط برای حذف این آلاینده‌ها است. در این پژوهش، توان تجزیه نفت خام توسط باکتری Methylorubrum sp. در دو وضعیت سلول آزاد و تثبیت‌شده در آلژینات سدیم بررسی و فعالیت آنزیم‌های آلکان‌مونو‌اکسیژناز، سیتوکروم P450 و لیپاز به‌عنوان آنزیم‌های کلیدی در تجزیه ترکیبات نفتی ارزیابی شد. نتایج نشان داد که در حضور چهار درصد نفت خام (۳۲۰۰ میلی‌گرم در لیتر)، باکتری در حالت آزاد موفق به تجزیه حدود 50درصد از ترکیبات نفتی شد، درحالی‌که تثبیت آن در دانه‌های آلژیناتی موجب افزایش تجزیه به 70درصد شد. آنالیز GC-MS نشان داد آلکان‌های سبک (C4–C9) به‌طور کامل و ترکیبات سنگین (C14–C28) تا ۷۴ درصد در حالت تثبیت‌شده تجزیه شدند. تصاویر میکروسکوپی FE-SEM ساختار متخلخل دانه‌ها و توزیع مناسب سلول‌ها را نشان داد. بیشینه فعالیت آنزیمی آلکان‌مونو‌اکسیژناز، سیتوکروم P450 و لیپاز در سلول‌های تثبیت‌شده در روز سوم به ترتیب برابر با 9، 5/9 و 55/13 واحد بر میلی‌گرم بود. تثبیت باکتری موجب افزایش پایداری، حفاظت سلولی و ارتقای عملکرد تجزیه‌ای شد. نتایج این مطالعه نقش اثربخش تثبیت Methylorubrum sp. را در زیست‌پالایی نفت خام تأیید کرده و می‌تواند گامی مؤثر در توسعه سامانه‌های مقیاس‌پذیر، ایمن و دوستدار محیط‌زیست برای مدیریت آلودگی‌های نفتی به‌شمار رود.
Crude oil spills and the persistence of hydrocarbon compounds in the environment pose significant threats to both human health and ecological integrity. These pollutants—due to their toxicity, chemical stability, and tendency to accumulate in water and soil—disrupt trophic interactions and exacerbate environmental degradation. Bioremediation using microorganisms offers an effective, cost-efficient, and environmentally sustainable strategy for mitigating such contamination. In this study, the crude oil degradation capacity of Methylorubrum sp. was evaluated under both free-living and sodium alginate–immobilized conditions. The activities of three key enzymes involved in hydrocarbon catabolism—alkane monooxygenase, cytochrome P450, and lipase—were also assessed. Under exposure to 4% (3200 mg/L) crude oil, free cells degraded approximately 50% of the petroleum hydrocarbons, whereas alginate immobilization enhanced degradation efficiency to 70%. Gas chromatography–mass spectrometry (GC–MS) analysis demonstrated complete degradation of light n-alkanes (C4–C9) and up to 74% degradation of long-chain n-alkanes (C14–C28) in the immobilized treatment. Field emission scanning electron microscopy (FE-SEM) confirmed the porous architecture of the alginate beads and uniform entrapment of bacterial cells. On day three, the immobilized cells exhibited peak specific activities of alkane monooxygenase, cytochrome P450, and lipase at 13.55, 9.5, and 9.0 U/mg protein, respectively. Overall, immobilization improved microbial stability, conferred resistance to environmental stress, and significantly enhanced crude oil biodegradation. These findings demonstrate the potential of immobilized Methylorubrum sp. for effective crude oil bioremediation and represent a promising step toward the development of scalable, safe, and environmentally responsible approaches to managing petroleum pollution.
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