Table of Content: 6 May, 2015; 14(18)  2015 May  A review on marine based nanoparticles and their potential applications DOI: 10.5897/AJB2015.14527[Article Number: 8AB382952706] Chinnappan Ravinder Singh, Kandasamy Kathiresan and Sekar Anandhan AbstractFull-Text PDF The increasing demands on nanoparticles have wide pertinent in almost all the fields. Marine ecosystem has variety of living resources, which includes prokaryotes like microorganism to eukaryotic organism like higher plants and animals. The present review dealt with the application of marine organisms in nanotechnology. Our discussion mainly focused on what the marine organisms are involved in and what type of... Read more.  2015 May  The use of multiplexed simple sequence repeat (SSR) markers for analysis of genetic diversity in African rice genotypes DOI: 10.5897/AJB2015.14478[Article Number: 9E756EA52708] Bonny M. Oloka, , Jimmy Lamo, Patrick Rubaihayo, Paul Gibson and Juan Vorster AbstractFull-Text PDF Rice is an emerging food and cash crop in Eastern Africa. Thousands of germplasm accessions have been introduced from major rice breeding centers, such as the International Rice Research Institute (IRRI), and Africa Rice but the genetic variability among the introduced rice germplasm is unknown. Knowledge on genetic diversity would be useful in designing measures for comprehensive breeding and conservation. To address... Read more.  2015 May  Molecular diversity study of black cumin (Nigella sativa L.) from Ethiopia as revealed by inter simple sequence repeat (ISSR) markers DOI: 10.5897/AJB2015.14567[Article Number: FC7397852709] Birhanu Kapital, Tileye Feyissa, Yohannes Petros and Said Mohammed AbstractFull-Text PDF http://www.academicjournals.org/journal/AJB/article-abstract/8AB382952706 http://www.academicjournals.org/journal/AJB/article-abstract/8AB382952706 http://www.academicjournals.org/journal/AJB/article-abstract/8AB382952706 http://www.academicjournals.org/journal/AJB/article-abstract/8AB382952706 http://www.academicjournals.org/journal/AJB/article-abstract/9E756EA52708 http://www.academicjournals.org/journal/AJB/article-abstract/9E756EA52708 http://www.academicjournals.org/journal/AJB/article-abstract/9E756EA52708 http://www.academicjournals.org/journal/AJB/article-abstract/9E756EA52708 http://www.academicjournals.org/journal/AJB/article-abstract/9E756EA52708 http://www.academicjournals.org/journal/AJB/article-abstract/FC7397852709 http://www.academicjournals.org/journal/AJB/article-abstract/FC7397852709 http://www.academicjournals.org/journal/AJB/article-abstract/FC7397852709 http://www.academicjournals.org/journal/AJB/article-abstract/FC7397852709 Nigella sativa L. (commonly known as black cumin) belonging to family Rannunculaceae is an important medicinal plant with worldwide distribution. In Ethiopia, N. sativa occurs in all regions and agro-ecologies at different altitudinal ranges. This plant has a lot of importance in Ethiopia. However, there is no information available on molecular genetic diversity of this crop in respect to Ethiopia. Therefore, the aim of... Read more.  2015 May  Inter simple sequence repeat (ISSR) analysis of Ethiopian white lupine (Lupinus albus L.) DOI: 10.5897/AJB2014.14379[Article Number: A050C1F52710] Abdie Oumer, Petros Yohannes, Tesfaye Kassahun, Teshome Abel and Bekele Endashaw AbstractFull-Text PDF White lupine (Lupinus albus L.) collected from two zones (West Gojjam and Awi) of Amhara region and one zone (Metekel) of Benishangul - Gumuz regional state of Ethiopia were studied using inter simple sequence repeat (ISSR) markers in an attempt to assess the genetic diversity. Four ISSR primers of which three were dinucleotide repeats and one, a penta nucleotide repeat amplified a total of 39 clear and reproducible... Read more.  2015 May  Genotype x environment interaction and stability analysis for yield and yield related traits of Kabuli-type Chickpea (Cicer arietinum L.) in Ethiopia DOI: 10.5897/AJB2014.14320[Article Number: EB941A052712] Getachew Tilahun Firew Mekbib, Asnake Fikre and Million Eshete AbstractFull-Text PDF Chickpea is the major pulse crop cultivated in Ethiopia. However, its production is constrained due to genotype instability and environmental variability. This research was carried out to examine the magnitude of environmental effect on yield of chickpea genotypes and to investigate the stability and adaptability of genotypes under different agro-ecologies. Seventeen (17) genotypes were evaluated in randomized complete... Read more.  2015 May http://www.academicjournals.org/journal/AJB/article-abstract/FC7397852709 http://www.academicjournals.org/journal/AJB/article-abstract/A050C1F52710 http://www.academicjournals.org/journal/AJB/article-abstract/A050C1F52710 http://www.academicjournals.org/journal/AJB/article-abstract/A050C1F52710 http://www.academicjournals.org/journal/AJB/article-abstract/A050C1F52710 http://www.academicjournals.org/journal/AJB/article-abstract/A050C1F52710 http://www.academicjournals.org/journal/AJB/article-abstract/EB941A052712 http://www.academicjournals.org/journal/AJB/article-abstract/EB941A052712 http://www.academicjournals.org/journal/AJB/article-abstract/EB941A052712 http://www.academicjournals.org/journal/AJB/article-abstract/EB941A052712 http://www.academicjournals.org/journal/AJB/article-abstract/EB941A052712 http://www.academicjournals.org/journal/AJB/article-abstract/EB941A052712  Effects of the humic acid extracted from vermicompost on the germination and initial growth of Brachiaria brizantha cv. MG5 DOI: 10.5897/AJB2015.14443[Article Number: C69E66352713] Mariá Moraes Amorim, Henrique Duarte Vieira, Isabela Moraes Amorim, Leonardo Barros Dobbss, Bruno Borges Deminicis and Priscilla Brites Xavier AbstractFull-Text PDF The biological effects of humic substances on vegetables depend on the source of extraction and the concentration used, on the vegetable species and on the age of the plant. This study aimed to evaluate the effect of different humic acid (HA) doses extracted from vermicompost on the germination and initial growth of Brachiaria brizantha cv. MG5. To that end, germination tests were conducted in germination agents, as... Read more.  2015 May  Effect of alkaline treatment on the sulfate content and quality of semi- refined carrageenan prepared from seaweed Kappaphycus alvarezii Doty (Doty) farmed in Indian waters DOI: 10.5897/AJB2014.14203[Article Number: E38190A52714] J. Moses, R. Anandhakumar and M. Shanmugam AbstractFull-Text PDF Seaweed Kappaphycus alvarezii previously known as Eucheuma cottonii is one of the best sources of kappa carrageenan and is cultivated in Philippines, Indonesia, Malaysia and other countries including India. In the present study, semi-refined carrageenan (SRC) was prepared from K. alvarezii with different concentrations of KOH (6, 12, 18 and 24%) at 80 ± 2°C for 2 h; its sulfate contents were... Read more.  2015 May  Modification of chitin as substrates for chitinase DOI: 10.5897/AJB2014.14178[Article Number: 6BAD6FC52716] Nuniek Herdyastuti, Sari Edi Cahyaningrum, Mizan Tamimi and Adi Wirawan AbstractFull-Text PDF Enzymes are able to bind to their substrates specifically at the active site. The proximity and orientation of the substrates strongly increase the likelihood that productive E–S complexes will arise. Treated chitin (powder or flake) is more efficient than crystalline chitin. This is because http://www.academicjournals.org/journal/AJB/article-abstract/C69E66352713 http://www.academicjournals.org/journal/AJB/article-abstract/C69E66352713 http://www.academicjournals.org/journal/AJB/article-abstract/C69E66352713 http://www.academicjournals.org/journal/AJB/article-abstract/C69E66352713 http://www.academicjournals.org/journal/AJB/article-abstract/C69E66352713 http://www.academicjournals.org/journal/AJB/article-abstract/E38190A52714 http://www.academicjournals.org/journal/AJB/article-abstract/E38190A52714 http://www.academicjournals.org/journal/AJB/article-abstract/E38190A52714 http://www.academicjournals.org/journal/AJB/article-abstract/E38190A52714 http://www.academicjournals.org/journal/AJB/article-abstract/E38190A52714 http://www.academicjournals.org/journal/AJB/article-abstract/E38190A52714 http://www.academicjournals.org/journal/AJB/article-abstract/6BAD6FC52716 http://www.academicjournals.org/journal/AJB/article-abstract/6BAD6FC52716 http://www.academicjournals.org/journal/AJB/article-abstract/6BAD6FC52716 the latter is less active due to its insolubility. The structure of treated chitin is opened; this facilitates its... Read more.  2015 May  Isolation of microalgae species from arid environments and evaluation of their potentials for biodiesel production DOI: 10.5897/AJB2014.14327[Article Number: A822FE552717] Innocent Okonkwo Ogbonna and James Chukwuma Ogbonna AbstractFull-Text PDF Twenty-five (25) strains of microalgae were isolated and screened for growth, lipid accumulation and biodiesel production from arid environments of North East Nigeria. Isolates that produced biomass concentration (≥ 1.50 g L-1 cell dry weight), accumulated high concentrations of lipids (≥ 18% of the cell biomass) and could be purified on agar plates were selected for further studies. Four strains morphologically... Read more. http://www.academicjournals.org/journal/AJB/article-abstract/6BAD6FC52716 http://www.academicjournals.org/journal/AJB/article-abstract/A822FE552717 http://www.academicjournals.org/journal/AJB/article-abstract/A822FE552717 http://www.academicjournals.org/journal/AJB/article-abstract/A822FE552717 http://www.academicjournals.org/journal/AJB/article-abstract/A822FE552717 http://www.academicjournals.org/journal/AJB/article-abstract/A822FE552717 Quick Links Google Scholar h5-index: 35 SNIP indicator: 0.90 SCImago JR: 0.26 Abbreviation: Afr. J. Biotechnol. Language: English ISSN: 1684-5315 DOI: 10.5897/AJB Start Year: 2002 Published Articles: 11323 Editors - AJB Editor-In-Chief Dr. George Nkem Ude Department of Natural Sciences Bowie State University Bowie, MD USA. Dr. N. John Tonukari Department of Biochemistry Delta State University Abraka, Nigeria. Editorial Board Members Dr. Gunjan Mukherjee Agharkar Research Institute (ARI), Autonomous Institute of the Department of Science and Technology (DST) Government of India Pune, India. Prof. Dr. A.E. Aboulata Plant Pathology Research Institute (ARC) Giza, Egypt. Dr. S. K. Das Department of Applied Chemistry and Biotechnology University of Fukui Japan. Prof. A. I. Okoh https://scholar.google.com/citations?hl=en&view_op=list_hcore&venue=eXaqDR5mwc4J.2015 https://scholar.google.com/citations?hl=en&view_op=list_hcore&venue=eXaqDR5mwc4J.2015 http://www.journalindicators.com/indicators/journal/14940 http://www.journalindicators.com/indicators/journal/14940 http://www.scimagojr.com/journalsearch.php?q=14940&tip=sid&clean=0 http://www.scimagojr.com/journalsearch.php?q=14940&tip=sid&clean=0 http://www.scopus.com/authid/detail.url?authorId=6601907865 http://www.scopus.com/authid/detail.url?authorId=6602269718 Applied and Environmental Microbiology Research Group (AEMREG) Department of Biochemistry and Microbiology University of Fort Hare Alice, South Africa. Dr. Ismail Turkoglu Department of Biology Education Education Faculty Fırat University Elazığ, Turkey. Prof. T. K. Raja Department of Biotechnology PSG College of Technology (Autonomous) Coimbatore India. Dr. George Edward Mamati Horticulture Department Jomo Kenyatta University of Agriculture and Technology Nairobi Kenya. Prof. Sagadevan G. Mundree Department of Molecular and Cell Biology University of Cape Town Rondebosch, South Africa. Dr. Amlan Patra Department of Animal Nutrition West Bengal University of Animal and Fishery Sciences India. Dr. Maria J. Poblaciones Department of Agronomy and Forest Environment Engineering Extremadura University, Spain. Dr. Chong Wang College of Animal Science Zhejiang A&F University China. Dr. Martin Fregene Centro Internacional de Agricultura Tropical (CIAT) Cali, Colombia. Prof. O. A. Ogunseitan Laboratory for Molecular Ecology Department of Environmental Analysis and Design University of California, Irvine, CA USA. Dr. Ibrahima Ndoye UCAD, Faculte des Sciences et Techniques Departement de Biologie Vegetale Laboratoire Commun de Microbiologie IRD/ISRA/UCAD Dakar, Senegal. Dr. Bamidele A. Iwalokun Biochemistry Department Lagos State University Nigeria. Dr. Jacob Hodeba Mignouna Virginia State University Agricultural Research Station Petersburg, VA USA Dr. Bright Agindotan Plant, Soil and Entomological Sciences Dept University of Idaho, Moscow, ID USA. Dr. A. P. Njukeng Département de Biologie Végétale Faculté des Sciences Université de Dschang Dschang, Cameroun. Dr. E. Olatunde Farombi Drug Metabolism and Toxicology Unit Department of Biochemistry University of Ibadan Ibadan, Nigeria. Dr. Stephen Bakiamoh Michigan Biotechnology Institute International Lansing, MI USA. Dr. N. A. Amusa Institute of Agricultural Research and Training Obafemi Awolowo University Moor Plantation Ibadan, Nigeria. Dr. Desouky Abd-El-Haleem Environmental Biotechnology Department Bioprocess Development Department Genetic Engineering and Biotechnology Research Institute (GEBRI) Mubarak City for Scientific Research and Technology Applications Alexandria, Egypt. Dr. Simeon Oloni Kotchoni Department of Plant Molecular Biology Institute of Botany University of Bonn Bonn, Germany. Dr. Eriola Betiku German Research Centre for Biotechnology Biochemical Engineering Division Braunschweig , Germany. Dr. Daniel Masiga International Centre of Insect Physiology and Ecology Nairobi, Kenya. Dr. Essam A. Zaki Genetic Engineering and Biotechnology Research Institute (GEBRI) Alexandria, Egypt. Dr. Alfred Dixon International Institute of Tropical Agriculture (IITA) Ibadan, Nigeria. Dr. Sankale Shompole Dept. of Microbiology, Molecular Biology and Biochemisty University of Idaho Moscow, ID USA. Dr. Mathew M. Abang Germplasm Program International Center for Agricultural Research in the Dry Areas (ICARDA) Aleppo, Syria. Dr. Solomon Olawale Odemuyiwa Pulmonary Research Group Department of Medicine Heritage Medical Research Centre University of Alberta Edmonton, Canada. Prof. Anna-Maria Botha-Oberholster Department of Genetics Forestry and Agricultural Biotechnology Institute Faculty of Agricultural and Natural Sciences University of Pretoria Pretoria, South Africa. Dr. O. U. Ezeronye Department of Biological Science Michael Okpara University of Agriculture Umudike, Nigeria. Dr. Joseph Hounhouigan Maître de Conférence Sciences et technologies des aliments Faculté des Sciences Agronomiques Université d'Abomey-Calavi Cotonou Bénin. Prof. Christine Rey Dept. of Molecular and Cell Biology University of the Witwatersand Johannesburg, South Africa. Dr. Kamel Ahmed Abd-Elsalam Molecular Markers Lab. (MML) Plant Pathology Research Institute (PPathRI) Agricultural Research Center Giza, Egypt. Dr. Jones Lemchi International Institute of Tropical Agriculture (IITA) Onne, Nigeria. Prof. Greg Blatch Department of Biochemistry, Microbiology & Biotechnology Rhodes University Grahamstown, South Africa. Dr. Jackie Hughes Research for Development International Institute of Tropical Agriculture (IITA) Ibadan, Nigeria. Dr. Robert L. Brown Southern Regional Research Center U.S. Department of Agriculture Agricultural Research Service New Orleans, LA USA. Dr. Deborah Rayfield Bowie State University Department of Natural Sciences Bowie, MD USA. Dr. Marlene Shehata University of Ottawa Heart Institute Genetics of Cardiovascular Diseases Ottawa, ON Canada. Dr. Hany Sayed Hafez The American University Cairo, Egypt. Dr. Clement O. Adebooye Department of Plant Science Obafemi Awolowo University, Ile-Ife, Nigeria. Dr. Ali Demir Sezer Marmara Üniversitesi Eczacilik Fakültesi Istanbul, Turkey. Dr. Anant B. Patel Centre for Cellular and Molecular Biology Hyderabad, India. Prof. Arne Elofsson Department of Biophysics, Biochemistry and Bioinformatics Stockholm University Sweden. Prof. Bahram Goliaei Departments of Biophysics and Bioinformatics Laboratory of Biophysics and Molecular Biology Institute of Biochemistry and Biophysics University of Tehran Tehran, Iran. Dr. Nora Babudri Dipartimento di Biologia cellulare e ambientale Università di Perugia Via Pascoli, Italy. Dr. S. Adesola Ajayi Seed Science Laboratory Department of Plant Science Faculty of Agriculture Obafemi Awolowo University Ile-Ife, Nigeria. Dr. Yee-Joo Tan Department of Microbiology Yong Loo Lin School of Medicine National University Health System (NUHS), National University of Singapore Singapore. Prof. Hidetaka Hori Laboratories of Food and Life Science Graduate School of Science and Technology Niigata University Niigata, Japan. Prof. Thomas R. DeGregori University of Houston Texas, USA. Dr. Wolfgang Ernst Bernhard Jelkmann Medical Faculty University of Lübeck Germany. Dr. Moktar Hamdi Department of Biochemical Engineering Laboratory of Ecology and Microbial Technology National Institute of Applied Sciences and Technology Tunisia. Dr. Salvador Ventura Department de Bioquímica i Biologia Molecular Institut de Biotecnologia i de Biomedicina Universitat Autònoma de Barcelona Bellaterra, Spain. Dr. Claudio A. Hetz Faculty of Medicine, University of Chile Santiago, Chile. Prof. Felix Dapare Dakora Research Development and Technology Promotion Cape Peninsula University of Technology Cape Town, South Africa. Dr. Geremew Bultosa Department of Food Science and Post harvest Technology Haramaya University Dire Dawa, Ethiopia. Prof. José Eduardo Garcia Conservation Genetics and Evolution Centro Acadêmico de Vitória Universidade Federal de Pernambuco - UFPE Rua Alto do Reservatório, s/n, Bela Vista, Vitória de Santo Antão-PE. CEP: 55608-680 Brasil Prof. Nirbhay Kumar Malaria Research Institute Department of Molecular Microbiology and Immunology Johns Hopkins Bloomberg School of Public Health Baltimore, MD USA. Prof. M. A. Awal Department of Anatomy and Histplogy Bangladesh Agricultural University Mymensingh, Bangladesh. Prof. Christian Zwieb Department of Molecular Biology University of Texas Health Science Center Tyler, TX USA. Prof. Danilo López-Hernández Instituto de Zoología Tropical, Facultad de Ciencias, Universidad Central de Venezuela. Institute of Research for the Development (IRD) Montpellier, France. Dr. Ekhaise Osaro Frederick University of Benin Faculty of Life Science Department of Microbiology Benin, Nigeria. Dr. Luísa Maria de Sousa Mesquita Pereira Institute of Molecular Pathology and Immunology at the University of Porto (IPATIMUP) Porto, Portugal. Dr. Min Lin Animal Diseases Research Institute Canadian Food Inspection Agency Ottawa, ON Canada. Prof. Nobuyoshi Shimizu Department of Molecular Biology Center for Genomic Medicine Keio University School of Medicine Tokyo, Japan. Dr. Adewunmi Babatunde Idowu Department of Biological Sciences University of Agriculture Abia, Nigeria. Dr. Yifan Dai Revivicor Inc. Pittsburgh, PA USA. Prof. Giuseppe Novelli Department of Biopathology Tor Vergata University Rome, Italy. Prof. Jean-Marc Sabatier Ingénierie des Peptides à Visée Thérapeutique, Université de la Méditerranée-Ambrilia Biopharma Inc. Faculté de Médecine Nord, Marseille, France. Dr. Fabian Hoti PneumoCarr Project Department of Vaccines National Public Health Institute Finland. Prof. Irina-Draga Caruntu Department of Histology Gr. T. Popa University of Medicine and Pharmacy Iasi, Romania. Dr. Dieudonné Nwaga Soil Microbiology Laboratory Biotechnology Center Plant Biology Department University of Yaoundé I Yaoundé, Cameroon. Dr. Gerardo Armando Aguado-Santacruz Biotechnology CINVESTAV-Unidad Irapuato Departamento Biotecnología Guanajuato, Mexico. Dr. Abdolkaim H. Chehregani Department of Biology Faculty of Science Bu-Ali Sina University Hamedan, Iran. Dr. Abir Adel Saad Department of Biotechnology Institute of Graduate Studies and Research Alexandria University Egypt. Dr. Azizul Baten Department of Statistics Shah Jalal University of Science and Technology Sylhet, Bangladesh. Dr. Bayden R. Wood Australian Synchrotron Program School of Chemistry Monash University Victoria, Australia. Dr. G. Reza Balali Department of Biology University of Isfahan Isfahan, Iran. Prof. H. Sunny Sun Institute of Molecular Medicine National Cheng Kung University Medical College Tainan, Taiwan. (R.O.C.) Prof. Ima Nirwana Soelaiman Department of Pharmacology Faculty of Medicine Universiti Kebangsaan Malaysia Kuala Lumpur, Malaysia. Prof. Tunde Ogunsanwo Faculty of Science Olabisi Onabanjo University Ago-Iwoye, Nigeria. Dr. Evans C. Egwim Federal Polytechnic Bida Science Laboratory Technology Department Bida, Nigeria. Prof. George N. Goulielmos Medical School University of Crete Crete, Greece. Dr. Uttam Krishna Cadila Pharmaceuticals Limited Gujarat, India. Prof. Mohamed Attia El-Tayeb Ibrahim Botany Department Faculty of Science South Valley University Qena, Egypt. Dr. Nelson K. Ojijo Olang’o Department of Food Science & Technology Jomo Kenyatta University of Agriculture and Technology (JKUAT) Nairobi, Kenya. Dr. Pablo Marco Veras Peixoto University of New York NYU College of Dentistry New York, NY USA. Prof. T. E. Cloete Department of Microbiology and Plant Pathology University of Pretoria Pretoria, South Africa. Prof. Djamel Saidi Laboratoire de Physiologie de la Nutrition et de Sécurité Alimentaire Département de Biologie Faculté des Sciences Université d’Oran Algeria. Dr. Ulises Urzúa Faculty of Medicine, University of Chile Santiago, Chile. Dr. Aritua Valentine National Agricultural Biotechnology Center Kawanda Agricultural Research Institute (KARI) Kampala, Uganda. Prof. Viroj Wiwanitkit Department of Laboratory Medicine Faculty of Medicine Chulalongkorn University Bangkok, Thailand. Dr. Thomas Silou University of Brazzaville Congo. Prof. Burtram Clinton Fielding University of the Western Cape Western Cape, South Africa. Dr. Meltem Sesli College of Tobacco Expertise Celal Bayar University Manisa, Turkey. Dr. Idress Hamad Attitalla Omar El-Mukhtar University Faculty of Science, Botany Department El-Beida, Libya. Dr. Linga R. Gutha Washington State University Prosser, WA USA. Dr. Vipul Gohel DuPont Industrial Biosciences Danisco (India) Pvt Ltd Haryana India. Dr. Sang-Han Lee Department of Food Science & Biotechnology Kyungpook National University Daegu, Korea. Dr. Bhaskar Dutta Biotechnology High Performance Computing Software Applications Institute (BHSAI) U.S. Army Medical Research and Materiel Command Frederick, MD USA. Dr. Muhammad Akram Faculty of Eastern Medicine and Surgery Hamdard Al-Majeed College of Eastern Medicine Hamdard University Karachi. Dr. M. Muruganandam Department of Biotechnology St. Michael College of Engineering & Technology Kalayarkoil, India. Dr. Gökhan Aydin Suleyman Demirel University Atabey Vocational School Isparta Türkiye. Dr. Rajib Roychowdhury Centre for Biotechnology (CBT) Visva Bharati, India. Dr. Takuji Ohyama Faculty of Agriculture Niigata University Niigata, Japan. Dr. Mehdi Vasfi Marandi University of Tehran Iran. Dr. Fügen Durlu-Özkaya Gazi Üniversity Dept. of Gastronomy and Culinary Art Ankara, Turkey. Dr. Reza Yari Islamic Azad University Tehran, Iran. Dr. Zahra Tahmasebi Fard Islamic Azad University Tehran, Iran. Dr Ping Zheng Zhejiang University Hangzhou China. Dr. Kgomotso P. Sibeko University of Pretoria South Africa. Dr. Greg Spear Rush University Medical Center Chicago, IL USA. Prof. Pilar Morata University of Malaga Malaga, Spain. Dr. Jian Wu Harbin Medical University China. Dr. Hsiu-Chi Cheng National Cheng Kung University and Hospital Taiwan. (R.O.C.) Prof. Pavel Kalac University of South Bohemia Czech Republic Dr Kürsat Korkmaz Ordu University Faculty of Agriculture Department of Soil Science and Plant nutrition Ordu, Turkey. Dr. Shuyang Yu Department of Microbiology University of Iowa Iowa City, IA USA. Dr. Mousavi Khaneghah College of Applied Science and Technology Department of Applied Food Science Tehran, Iran. Dr. Qing Zhou Department of Biochemistry and Molecular Biology Oregon Health and Sciences University Portland Portland, OR USA. Dr. Legesse Adane Bahiru Department of Chemistry Jimma University Ethiopia. Dr. James John School of Life Sciences Pondicherry University Kalapet, India. Dr. Ramesh B Narasingappa Division of Biotechnology University of Agricultural Sciences (UAS) Agricultural College Bangalore, India. Dr. Prabhat Kumar Singhal Center for Computational and Integrative Biology (CCIB) Department of Genetic Massachusetts General Hospital/Harvard Medical School USA. Dr. Muruganantham Mookkan Division of Plant Sciences University of Missouri Columbia, USA. Dr. Lixia Zhao Cell and Molecular Physiology Loyola University Medical Center USA. Dr. Elsayed Ziedan National Research Centre Plant Pathology Department Dokki, Egypt. Prof. Hazim Jabbar Al-Daraji University of Baghdad College of Agriculture Baghdad, Iraq. Dr. Vijay Kumar Garlapati Department of Biotechnology and Bioinformatics Jaypee University of Information Technology Waknaghat, India. Dr. Harsh Kavi Albert Einstein College of Medicine Bronx, NY USA. Dr. Berhanu Abraha Biology Department Bahir Dar Univesity Ethiopia. Dr. Fazal Shirazi Infectious Disease Department The University of Texas MD. Anderson Cancer Center USA. Dr. Yi Zhu Internal Medicine Deparment UT Southwestern Medical Center Dallas, USA. Dr. Wen-Li Du National Heart, Lung, and Blood Institute National Institutes of Health (NIH) Bethesda, USA. Dr. Lei Wang Max Planck Institute for Heart and Lung Research Dept. of Pharmacology Bad Nauheim, Germany. Dr. Shrawan Mishra The Hormel Institute University of Minnesota Austin, MN USA. Dr. Dipnarayan Saha Biotechnology Unit, Division of Crop Improvement Central Research Institute for Jute and Allied Fibres (Indian Council of Agricultural Research) Barrackpore, India. Dr. Imad Hadi Babylon University Faculty of Science Biotechnology Department Iraq. Dr. Vincenzo Tufarelli Department of Emergency and Organ Transplant (DETO) Section of Veterinary Science and Animal Production University of Bari “Aldo Moro” Italy. Dr. Carmelo Peter Bonsignore Department PAU – Laboratorio di Entomologia ed Ecologia Applicata Mediterranean University of Reggio Calabria Italy. Dr. Christophe Brugidou Research Institute for Development (IRD) Center France. Dr. Preejith Vachali School of Medicine University of Utah USA. Dr. Balabhadrapatruni V.S.K. Chakravarthi Michigan Center for Translational Pathology Dept of Pathology University of Michigan Ann Arbor, USA. Dr. Srećko Trifunović Department of Chemistry Faculty of Science University of Kragujevac Serbia. Dr. Huda El-Sheshtawy Biotechnological Application lab., Process, Design and Development Egyptian Petroleum Research Institute (EPRI) Cairo, Egypt. Dr. Sekhar Kambakam Department of Agronomy Iowa State University USA. Dr. Desobgo Zangue Steve Carly Food Processing and Quality Control University Institute of Technology (University of Ngaoundere) Cameroon. Dr. Girish Kamble Botany Department SRRL Science College Morshi India. Dr. Zhiguo Li School of Chemical Engineering University of Birmingham UK. Dr. Alok Pandey Pharmacology and Physiology New Jersey Medical School Rutgers University, USA. Dr. Ajit Waman Division of Horticulture and Forestry ICAR- Central Island Agricultural Research Institute Port Blair, India. Dr. Bidyut Saha Chemistry Department Burdwan University WB, India. Dr. Karthik Rajendran Department of Resource Recovery University of Boras Sweden. Dr. Rachana Bhatt New Jersey Institute of Technology Newark, NJ, USA. Dr. Ashraf El-Kereamy Agriculture and Natural Resources (ANR) University of California, USA. Dr. Ram Prasad Amity Institute of Microbial Technology Amity University India. Dr. Vijay Kumar Eedunuri Department of Cellular And Structural Biology UT health Science Center San Antonio, USA. Dr. Mahmoud Mohammed Department of Food Hygiene Faculty of Veterinary Medicine Mansoura University Egypt. Dr. K.V. Madhusudhan Botany Department Government College Kurnool, India. Prof. Naziha Hassanein Department of Microbiology Faculty of Science Ain Shams University Egypt. Dr. Shashi Kumar Synthetic Biology and Biofuels Institution International Center for Genetic Engineering and Biotechnology India. Dr. S. Ahmed John Department of Botany Jamal Mohamed College (autonomous) Tiruchirappalli, India. Dr. Jitendra Kumar Saini Indian Oil Corporation Ltd. Research & Development Centre Faridabad, India. 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http://www.academicjournals.org/AJB African Journal of Biotechnology Full Length Research Paper Modification of chitin as substrates for chitinase Nuniek Herdyastuti*, Sari Edi Cahyaningrum, Mizan Tamimi and Adi Wirawan Department of Chemistry, Surabaya State University, Jl. Ketintang Surabaya, Indonesia. Received 11 September, 2014; Accepted 27 April, 2015 Enzymes are able to bind to their substrates specifically at the active site. The proximity and orientation of the substrates strongly increase the likelihood that productive E–S complexes will arise. Treated chitin (powder or flake) is more efficient than crystalline chitin. This is because the latter is less active due to its insolubility. The structure of treated chitin is opened; this facilitates its interaction with the enzyme. The purpose of this research was to create a kind of modified chitin and study the characterization of the different types of chitin including functional groups by IR spectrophotometer, pore size, surface area and crystallinity by X-Ray diffraction. Chitin from shrimp shell was modified into colloidal, bead, amorphous and superfine chitin. The results of the IR spectra of colloidal and bead chitin showed a similar pattern with chitin powder; they peaked at 3447 and 3113 cm -1 (OH and NH2 groups), 1645 cm -1 (amide groups N-H) and 1071 cm -1 (group C-O). Superfine and amorphous chitin had similar absorbance with powder chitin but appeared to peak in the fingerprint region. Characterization of physical properties based on the pore size and surface area of powder, colloidal, superfine, amorphous and bead chitin changed the pore radius of each type of chitin due to the treatment of swelling. Crystallinity showed that specific diffractogram pattern in the three main peaks 2 was 9.5, 19.5 and 26 with varying intensity. Chitinase activity assay using modified types of chitin substrate had higher values than chitin powder. The highest activity was in amorphous chitin with values of 1.858 U/mL. This is because it has chitin chain and the rearrangement of its structure was more open, facilitating its interaction with enzyme. Key words: Chitin modified, chitinase, substrate. INTRODUCTION Chitin is a polymer that is very abundant in nature and is second only to cellulose. It is widely spread in nature as in fungi, algae, nematodes, arthropods, molluscs, plants and animals (Guo et al., 2004). So far, it has been found that it has very little large-scale industrial use because of its extreme insolubility; it cannot be absorbed or digested directly in the gastrointestinal tract (Dai, 2011). Chitin structure has three forms, namely α, β and γ. - Chitin is a form of a dense structure isomorphous having strong hydrogen bonds. -Chitin has a structure with weaker intramolecular bonds but slightly more stable than α-chitin. -Chitin is a combination of α and β chitin structure. -Chitin structure causes the chitin not to be soluble in the solvent, while β-chitin can be swollen in water, as *Corresponding author. E-mail: nuniekherdyastuti@gmail.com. Abbreviations: SDS, Sodium dodecyl sulfate; Ct, chitin powder; Cc, colloidal chitin; Sf, superfine; Cb, bead; Ca, amorphous. Author(s) agree that this article remains permanently open access under the terms of the Creative Commons Attribution License 4.0 International License http://www.academicjournals.org/AJB http://creativecommons.org/licenses/by/4.0/deed.en_US http://creativecommons.org/licenses/by/4.0/deed.en_US chitin is soluble in formic acid (Coutinõ et al., 2006). Chitin chains between each other are associated with very strong hydrogen bonds between the NH groups of one chain and the C = O groups of adjacent chains. Hydrogen bonds make chitin to be insoluble in water and to form fiber (Rostinawati, 2008). The presence of chitin in nature varies abundantly and degrades rapidly, due to the presence of some bacteria and fungi that have chitinase enzyme capable of degrading chitin. Chitin can be degraded in two lines: the first is a degradation by mechanisms that hydrolyze bonds chitinolytic 1,4-ß- glycosides, or polymers having first deacetylation and then hydrolysed by chitosanase (Herdyastuti et al., 2009). Chitinases are extracellular inducible enzymes that catalyze the first step in chitin digestion, hydrolysis of - 1,4 linkages between the N-acetyl glucosamine (NAG) molecules. They are found in a variety of organisms including viruses, bacteria, fungi insects, higher plants and animals and play important physiological roles depending on their origin (Kuddus and Ahmad, 2013). Chitinolytic activity induces strain in the growth medium in the presence of chitin as a carbon source (Chernin et al., 1998). Chitin can be modified by adding acid, base or detergents such as sodium dodecyl sulfate (SDS) for it to become swollen (Illankovan et al., 2007). The swelling of chitin powder process is expected to help the enzymatic reaction between chitinase and chitin. Possible modification of chitin-chitin can cause structural changes to become more open due to the restructuring of the chitin-chain. Possible rearrangement of chitin structure can cause changes in the functional group or the physical properties of each type of chitin compared with chitin powder. MATERIALS AND METHODS Preparation of chitin Chitin was obtained from shrimp shells that have been dried and pulverized and the isolation was done by the method of Acosta et al. (1993). Chitin isolation process consists of two stages: deproteinisation and demineralization. Chitin is made into the form of colloids according to Hsu and Lockwood (1975). Chitin was dissolved in concentrated HCl (37%), and then precipitated as a colloidal suspension with the addition of cold water (5°C). The suspension was filtered and the residue was washed with distilled water until it got to neutral pH, and then dried with an oven. This process gives ± 85% recovery. Chitin was then prepared into colloidal, superfine, bead and amorphous form. In the colloidal form, chitin is made by adding concentrated HCl (37%). Chitin beads were obtained by dissolving in 2% formic acid and 2 M NaOH solution. Chitin amorphous was prepared by dissolving chitin in a mixed solution of 40% NaOH and 0.2% SDS (which has been cooled to a temperature of 4°C). Solution was in-swell for 1 h at 4°C and matrix slurry was stored for 1 night at -20°C temperature, and then neutralized with HCl 6 N. Furthermore, it was filtered and washed with ethanol, water and acetone. The result was dried with a freeze dryer. Herdyastuti et al. 1591 Produce of chitinase enzyme Chitinase was produced in medium with the following composition: 0.4% chitin, 0.7% K2HPO4, 0.3% KH2PO4, 0.5% MgSO4.5H2O, 0.01% FeSO4.7H2O, 0.001% MnCl2 and 0.5% peptone, and incubated at room temperature for 45 h in rotary shaker at 150 rpm. The culture cells were centrifuged at 4000 rpm for 20 min (4°C). The supernatant was brought to 50% saturation with ammonium sulphate at 4°C for 30 min by stirring magnetic stirrer. The precipitate was recovered by centrifugation at 4000 rpm for 30 min (4°C) and pellet formed was solubilized in 0.1 M phosphate buffer pH 7.0. The solution was dialyzed overnight against the same buffer at 4°C. Chitinase assay Chitinase activity was measured by colorimetric method based on the released N-acetyl-glucosamine(Monreal and Reese, 1969). The colloidal chitin solution (2.0 mL of 1.25% (w/v)) dissolved in 200 mM potassium phosphate buffer was added to 0.5 mL enzymes solution and incubated for 2 h at room temperature. The suspensions were centrifuged at 4000 rpm for 10 min and then supernatant (1.0 mL) was added to 2.0 mL deionized water and 1.5 mL color reagent (5.3 M sodium potassium tartrate and 3,5-dinitrosalicyclic acid 96 mM). The mixed solution was placed in boiling water for 5 min and cooled at room temperature, and then the absorbance was measured at 540 nm. One unit (U) of chitinase activity was defined as the amount of enzyme required to release 1.0 mg N-acetyl D- glucosamine from chitin per hour. Characterization of substrates The structure of substrates was determined by FT-IR spectro- photometer (Perkin Elmer); the analysis of pore size and surface area was done with high speed surface area (NOVA 1200e). X-ray diffractograms were recorded by a Bruker type D 8 advance. RESULTS AND DISCUSSION Characteristic of chitin Chitin that has been isolated from waste shrimp shells chitin powder (Ct) and has been modified into a kind of colloidal chitin (Cc), Superfine (Sf), bead (Cb) and amorphous (Ca) is as shown in Figure 1. Chitin was modified to have almost the same colour, smoother texture and lighter but more types of colloidal tawny color and form larger granules. The yield of the average obtained was 50 to 60%. The results of the analysis of the functional group on IR- spectrophotometer modified chitin (Figure 2) show the absorption at 3446 and 3113 cm -1 (OH and NH2 groups). Sharp absorption peaked at 1645 cm -1 indicates the presence of amide groups (N - H) and 1071 cm -1 shows the group C - O. The spectra of Ca are a sharp peak in the fingerprint region below 700 cm -1 which is not found in other types of chitin. Spectra of superfine showed similarity with the IR spectra of chitin and colloidal chitin. The characteristic peaks of chitin are -OH group (3433 cm -1 ), -NH (amide) at 1587.54 cm -1 , CH bending vibration 1592 Afr. J. Biotechnol. Figure 1. (A) Powder chitin. (B) Colloidal chitin. (C) Superfine chitin. (D) Amorphous chitin. (E) Bead chitin. Figure 2. Spectra-IR of (A) Powder chitin, (B) Colloidal chitin, (C) Superfine chitin, (D) Bead chitin and (E) Amorphous chitin. modified into a kind of colloidal chitin (Cc), Superfine (Sf), bead (Cb) and amorphous (Ca) as shown in Figure 1. Figure 1. Chitin powder (A), Colloidal (B), Superfine (C), Bead (D) and Amorphous (E) Chitin that has been modified to have almost the same colour, smoother texture and lighter but B C D E A D E A B C Herdyastuti et al. 1593 Table 1. The relevant peak FT-IR spectra of chitin substrates. Function group Wave number (cm -1 ) Ct Cc Sf Cb Ca C – O 1379 1380 1379 1379 1308 N – H (bending) 1562 1635 1567 1567 - C – H (streching) 2886 2931 - - 2963 C = O 1651 1635 1633 1633 - O – H 3446 3446 3435 3435 3345 Ct, Powder chitin; Cc, colloidal chitin; Sf, superfine chitin; Ca, amorphous chitin; Cb, bead chitin. Table 2. Analysis of physical characteristic in the chitin substrates. Substrate Pore radius (A) Pore area (m 2 /g) Pore volume (cc/g) Powder chitin Colloidal chitin Superfine chitin 19.108 19.044 162.879 1.365 2.780 0.06 4.00 x 10 -2 5.00 x 10 -2 0,0001 Amorphous chitin 19.159 3.252 1.30 x 10 -2 Bead chitin 19.011 0.606 4.00 x 10 -3 at 1378.7 cm -1 , stretching vibration of C = O, amide - NHCOCH3 (1633.1 cm -1 ) and CO alcohol at 1072.9 cm -1 (Tamimi and Herdyastuti, 2013). Table 1 shows the relevant peak FT-IR spectra of chitin substrates. The other research shows that chitin has -NH peak at 3269, 1663 and 1629 cm -1 due to the reduction of primary amides that generate elimination of carboxyl groups (Coutin˜o et al., 2006). The results of the analysis based on the physical properties of pore size and surface area of powder, colloidal, superfine, amorphous and bead chitin are shown in Table 2. The results of the analysis showed that treatment of swelling on each type of chitin changed pore radius and became larger. Data show pore radius of superfine chitin is 9 times greater than the powder chitin. Wide pores of colloidal chitin also increased, but the chitin beads were apparently amorphous and their volume was reduced. The changes of volume size and pore radius would affect the interaction of enzymes with the substrate. Diffractogram of chitin powder, colloidal and bead shows the same pattern. There are 3 main peaks 2 of 9.5, 19.5 and 26; the intensity tends to be weaker in colloidal and bead chitin than in powder chitin as shown by the studies of Illankovan et al. (2007), in which the diffractogram of powder, colloidal and amorphous chitin had main peak of 2 of 9.4 and 20. This shows that colloidal and bead chitin has a lower degree of crystallinity than chitin powder. Swelling process in colloidal and bead chitin causes larger pores and is easily inflated in water medium; this leads to the easy interaction of enzymes with substrates than in the form of chitin powder, which is more compacted (Figure 3). Chitinase activity Chitin from crab shell is identified as the best carbon source and colloidal chitin is reported as the best source for producing chitinase. The results show the highest activity for the amorphous chitin after colloidal chitin types (Figure 4). Suraini et al. (2008) reported that the highest specific activity produced by colloidal chitin was 14.59 U/mg. Chitinase from Paenibacillus sp. D1 showed that the highest activity was 35 U/mL at 30°C after 72 h (Singh, 2010). The results of optimization of culture nutrients revealed that the amount of colloidal chitin as a sole carbon source in the growth medium of Trichoderma viride was 32.1 U/mL (Sharaf et al., 2012). By optimizing the above cultural conditions, the production of chitinase from Bacillus amyloliquefaciens SM3 increased by three fold to 33.5 U/mL at the final stage (Das et al., 2012). This form of chitin is tight due to its anti-parallel chain form and it stabilizes polymorphism shape naturally causing chitin not to dissolve in the solvent (Majtán et al., 2007). Modified chitin using SDS detergent leads to swelling of chitin structure causing changes in the physical properties of chitin powder. Amorphous type chitin has fingers longer than other types of chitin and three times larger area than the chitin powder. The data indicate that amorphous chitin is more open and more 1594 Afr. J. Biotechnol. Figure 3. Difractograme of powder chitin (A), colloidal chitin (B), amorphous chitin (C), bead chitin (D) and superfine chitin (E). Figure 4. Chitinase activity with substrate types. Ct, Powder chitin; Cc, colloidal chitin; Sf, Superfine chitin; Ca, amorphous chitin; Cb, bead chitin. A 2 Degree In te n si ty B C D E daDad Aasd likely to facilitate interaction with chitinase that can provide higher chitinase activity than other substrates. Ilankovan (2005) reported that among the chitinolytic activities of the commercial enzymes investigated with amorphous chitin as substrate, the bovine pepsin had the highest chitinolytic activity. Conclusion Modification of chitin by adding detergents causes characteristic changes in its physical properties and the structure becomes more open than chitin powder, thus causing its interaction with the enzyme chitinase. Amorphous chitin can be used as an alternative substrate or inducer for chitinase enzyme indicated by higher chitinase activity than using chitin powder. Conflict of interests The authors did not declare any conflict of interest. ACKNOWLEDGEMENTS The research was supported by Ditlitabmas, DP2M- DIKTI, National Education Department of Indonesia. 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