International Journal of Current Research and Review
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IJCRR - Vol 05 Issue 08, April, 2013

Pages: 11-19

Date of Publication: 30-Nov--0001


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IN VITRO EVALUATION OF AFLATOXIN M1 CONTROL POTENTIAL OF SIX ESSENTIAL OILS IN BENIN

Author: Philippe Sessou, Fidele Tchobo, Pascal Agbangnan, Elvis Adjalian, Paulin Azokpota, Issaka Youssao, Souaibou Farougou

Category: General Sciences

Abstract:Aflatoxin M1 (AFM1) is hepatocarcinogen frequently found in cheese. Its presence in this foodstuff constitutes a serious threat for consumers. The present study had evaluated aflatoxin M1 control potential of six essential oils against synthetic aflatoxin M1. It had consisted to blend Aflatoxin M1 diluted in acetonitrile (1\?g/mL) with essential oil to achieve final concentrations of 2%, 4%, 6%, 8% and 10% of oil and to follow the disintegration of this AFM1 in the meantime of 30 minutes, 1, 2, 3, 4, 5 and 24 hours on silica gel with or without thin layer chromatography assay (TLC). The results obtained revealed that at the concentrations tested, no fluorescence was observed for the mixtures of Aflatoxin M1 with C. zeylanicum, P. racemosa and Syzygium aromaticum essential oils on silica gel without chromatography assay contrary to results obtained for the same mixtures after chromatography assay. The mixture of Cymbopogon citratus showed fluorescence with or without TLC assay. Pure extracts of Ocimum gratissimum and Zingiber officinale showed a green fluorescence. In sum, all the essential oils tested didn't possess capacity to destroy AFM1 at concentrations investigated. Although, these extracts cannot be used for controlling aflatoxin M1, their potentiality for inhibiting mycotoxins production by moulds can be considered.

Keywords: Aflatoxin M1, antiaflatoxin assay, essential oils, TLC assay

Full Text:

INTRODUCTION
Benin is underdeveloped country which produces milk with an output of 94 million liters in 2009 (FAO, 2010). Milk produced in Benin, instead of domestic consumption, is processed to many dairy products among which traditional cheese wagashi is the most consumed largely by rural than citizens populations due to its content in good source of many nutrients such as proteins and calcium (Dossou et al., 2006; Keke et al., 2008). However, Benin country has favourable warm and humid climate, socio-economic (ignorance of the toxin and poor infrastructure to manage mycotoxins prevention strategies) and compelling environmental (drought) factors that enhance the growth of aflatoxigenic fungi and subsequently aflatoxins production inside wagashi (Okeke et al., 2012; Sessou et al., 2012a, b). Aflatoxins are toxic fungal metabolites produced by Aspergillus species, mainly by Aspergillus flavus and Aspergillus parasiticus, but also by Aspergillus nomius, Aspergillus pseudotamarii, Aspergillus ochraceus (IARC, 2012; Siddappa et al., 2012). Aflatoxins consist of a group of approximately 20 related metabolites, among which aflatoxins B1, B2, G1 and G2 are often found in foods. Aflatoxin B1 (AFB1) is metabolized by the animals consuming these contaminated feeds to AFM1 mainly by the hepatic microsomal mixed-function oxidase system (Siddappa et al., 2012). Aflatoxin M1 (AFM1) was classified by the International Agency for Research on Cancer (IARC) as a group 1 human carcinogen (IARC, 2012, Okeke et al., 2012). Thus, its potential risk to human health makes its presence in milk products such as cheese wagashi undesirable. AFM1 is unaffected by pasteurization and ultra-high-temperature (UHT) treatment (Tekinsen and Eken, 2008). Alternative methods for its control are needed to be performed. The control or disintegration of AFM1 by natural agents as essential oils, safe for human (Burt, 2004) in order to reduce or eliminate this toxic metabolite in wagashi eventually contaminated is necessary for minimizing public health hazards. The objective of this work was to evaluate aflatoxin M1 control potential of six essential oils extracted in Benin.

MATERIAL AND METHODS
Material was constituted of six essential oils extracted by hydrodistillation from Cinnamomum zeylanicum, Cymbopogon citratus, Ocimum gratissimum, Pimenta racemosa, Syzygium aromaticum and Zingiber officinale previously analyzed by GC/MS and GC/FID (table 1) and  Aflatoxin M1 ( 10 µg/mL in acetonitrile, Lot: LB 96767; 46319-U, quantity: 1 mL, Exp: Nov/2015; USA) purchased at SUPELCO  Analytical  society based at Unity State of America and diluted in acetonitrile to have final concentration of 1µg/mL which is used for the assay.

The method used in this study was based on that described by Adjou et al. (2012a, b, c and 2013). It had consisted to mix Aflatoxin M1 diluted in acetonitrile (1µg/mL) with essential oil to achieve final concentrations of 2%, 4%, 6%, 8% and 10% of oil and to follow the inhibition or disintegration of this metabolite (AFM1) in the meantime of 30 minutes, 1, 2, 3, 4, 5 and 24 hours on silica gel without or after thin layer chromatography assay (TLC). Five microliter of mixture (Aflatoxin + essential oil) or pure extract of essential was spotted and air dried on TLC plates (TLC Silica gel 60 F254, Merck, Germany) and observed under long wave (365 nm) UV and then developed in the solvents system comprising TEF (Toluene/ethylacetate/Formic acid, 5:4:1 v/v/v) and CAP (Chloroform/acetone/2-propanol, 85:15:20, v/v/v), air dried and observed under long wave (365 nm) UV.

RESULTS AND DISCUSSION
The present study had investigated the potentiality of six essential to destroy Aflatoxin M1 in perspective for their use to control mycotoxins produced eventually inside traditional cheese wagashi. The results obtained from this work are prsented in figures 1 to 5 and showed that these extracts studied didn’t possess capacity to decontaminate food contaminated by aflatoxin M1. In fact, results obtained from this study revealed that at the concentrations tested, no fluorescence was observed for the mixtures of Aflatoxin M1 with Cinnamomum zeylanicum, Pimenta racemosa and Syzygium aromaticum essential oils on silica gel without chromatography assay contrary to results obtained for the same mixtures after chromatography assay. Indeed, these extracts had masked the fluorescence of Aflatoxin M1 when mixed together and spotted on TLC plate and observed under UV light 365 nm without tthin layer chromatography assay (figure1). Based on theory of Adjou et al. (2012 a, b, c and 2013), we can conclude that our three extracts C. zeylanicum, P. racemosa and S. aromaticum possessed antiaflatoxin M1 potential. But this conclusion will be wrong when we consider the elution of these mixtures which showed the fluorescence at the same bright with the standard AFM1 spotted (figure 2 to 5). The masking of fluorescence of Aflatoxin M1 when mixed with extracts of C. zeylanicum, P. racemosa and S. aromaticum could be explained by a reaction of components of these oils with the fluorescence pole of Aflatoxin M1 which is not inevitably sinonymous of the destroying of the toxic metabolite. These extracts may possess capacity to inhibit production of mycotoxins produced by toxinogenic mouldsThe mixture of Cymbopogon citratus showed fluorescence either with or without TLC assay. This last pure oil (C. citratus) with no fluorescence when spotted on TLC plate had not capacity to mask the aflatoxin M1 fluorescence when mixed together with the toxic metabolite. Pure extracts of Ocimum gratissimum and Zingiber officinale spotted alone showed a green fluorescence on silica gel without TLC assay and didn’t allow appreciating their activity at this step (figure 1). The present study had shown that the works of Adjou et al. (2012a,b,c and 2013) concerning the appreciation of aflatoxigenic inhibition potential of extracts based on their ability to inhibit fluorescence of aflotoxins presents insufficiencies. In fact, these authors have wrongly exploited the method of Nguyen (2007) and Atanda et al. (2011) whose works were made not for evaluating the aflatoxin production inhibition by extracts but for the capacity of strains of Aspergillus to produce aflatoxins revealed by fluorescence of these metabolites under UV light. Our work showed that the inhibition of fluorescence of the metabolite is not inevitably synonymous of capacity of extract to inhibit the metabolite production of toxinogenic fungus which normally grows. Also, the appearance of fluorescence after observing the mixture of essential oil plus the metabolite in culture medium is not synonymous of incapacity of extract to possess antiaflatoxin inhibition potential. Indeed, our study showed that the pure extracts of O. gratissimum and Z. officinale spotted alone showed green fluorescence. As recommendation, before conclude that the extracts possess antiaflatoxin inhibition potential based on inhibition of fluorescence, the previous authors (Adjou et al) must complete their work by proceeding to the detection and the quantification of the aflatoxins in the whole culture medium composed of extract, toxinogenic fungus which had its metabolite fluorescence inhibited or not. As alternative methods to detected capacity of mycotoxins production inhibition by essential oils, that described by Singh et al. (2010), Prakash et al. (2012a, b, c) and Shukla et al. (2012) are scientifically efficient and appropriate. In sum, this study showed that essential oils of C. zeylanicum, C. citratus, O. gratissimum, P. racemosa, S. aromaticum and Z. officinale didn’t possess potential to destroy aflatoxin M1 when presented inside food. Also, it showed that the appreciation of inhibitory aflatoxin production potential of essential oils based on their ability to inhibit fluorescence of metabolites produced by toxinogenic moulds presents more insufficiencies.  

CONCLUSION

The work has assessed the potential of six essential oils to control aflatoxin M1 in perspective to their use to destroy this toxic metabolite inside cheese wagashi contaminated. Results obtained showed that these extracts had not potential to disintegrate AFM1 at concentrations investigated. However, these extract especially C. zeylanicum, P. racemosa and S. aromaticum can be considered as promising extract for mycotoxins production inhibition.

ACKNOWLEDGMENTS
The authors are grateful to University Council of Development (CUD) for its financial support. They are also thankful to Professor Dominique SOHOUNHLOUE for his scientific contributions. Authors acknowledge the great help received from the scholars whose articles cited and included in references of this manuscript. The authors are also grateful to authors / editors / publishers of all those articles, journals and books from where the literature for this article has been reviewed and discussed. Authors are grateful to IJCRR editorial board members and IJCRR team of reviewers who have helped to bring quality to this manuscript.

References:

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A Study by Amr Y. Zakaria et al. entitled "Single Nucleotide Polymorphisms of ATP-Binding Cassette Gene(ABCC3 rs4793665) affect High Dose Methotrexate-Induced Nephrotoxicity in Children with Osteosarcoma" is awarded Best Article for Vol 13 issue 19
A Study by Kholis Ernawati et al. entitled "The Utilization of Mobile-Based Information Technology in the Management of Dengue Fever in the Community Year 2019-2020: Systematic Review" is awarded Best Article for Vol 13 issue 18
A Study by Bhat Asifa et al. entitled "Efficacy of Modified Carbapenem Inactivation Method for Carbapenemase Detection and Comparative Evaluation with Polymerase Chain Reaction for the Identification of Carbapenemase Producing Klebsiella pneumonia Isolates" is awarded Best Article for Vol 13 issue 17
A Study by Gupta R. et al. entitled "A Clinical Study of Paediatric Tracheostomy: Our Experience in a Tertiary Care Hospital in North India" is awarded Best Article for Vol 13 issue 16
A Study by Chandran Anand et al. entitled "A Prospective Study on Assessment of Quality of Life of Patients Receiving Sorafenib for Hepatocellular Carcinoma" is awarded Best article for Vol 13 issue 15
A Study by Rosa PS et al. entitled "Emotional State Due to the Covid – 19 Pandemic in People Residing in a Vulnerable Area in North Lima" is awarded Best Article for Vol 13 issue 14
A Study by Suvarna Sunder J et al. entitled "Endodontic Revascularization of Necrotic Permanent Anterior Tooth with Platelet Rich Fibrin, Platelet Rich Plasma, and Blood Clot - A Comparative Study" is awarded Best Article for Vol 13 issue 13
A Study by Mona Isam Eldin Osman et al. entitled "Psychological Impact and Risk Factors of Sexual Abuse on Sudanese Children in Khartoum State" is awarded Best Article for Vol 13 issue 12
A Study by Khaw Ming Sheng & Sathiapriya Ramiah entitled "Web Based Suicide Prevention Application for Patients Suffering from Depression" is awarded Best Article for Vol 13 issue 11
A Study by Purushottam S. G. et al. entitled "Development of Fenofibrate Solid Dispersions for the Plausible Aqueous Solubility Augmentation of this BCS Class-II Drug" is awarded Best article for Vol 13 issue 10
A Study by Kumar S. et al. entitled "A Study on Clinical Spectrum, Laboratory Profile, Complications and Outcome of Pediatric Scrub Typhus Patients Admitted to an Intensive Care Unit from a Tertiary Care Hospital from Eastern India" is awarded Best Article for Vol 13 issue 09
A Study by Mardhiah Kamaruddin et al. entitled "The Pattern of Creatinine Clearance in Gestational and Chronic Hypertension Women from the Third Trimester to 12 Weeks Postpartum" is awarded Best Article for Vol 13 issue 08
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A Study by Kumari PM et al. entitled "Study to Evaluate the Adverse Drug Reactions in a Tertiary Care Teaching Hospital in Tamilnadu - A Cross-Sectional Study" is awarded Best Article for Vol 13 issue 05
A Study by Anu et al. entitled "Effectiveness of Cytological Scoring Systems for Evaluation of Breast Lesion Cytology with its Histopathological Correlation" is awarded Best Article of Vol 13 issue 04
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A Study by Mohita Ray et al. entitled "Accuracy of Intra-Operative Frozen Section Consultation of Gastrointestinal Biopsy Samples in Correlation with the Final Histopathological Diagnosis" is awarded Best Article for Vol 13 issue 01
A Study by Badritdinova MN et al. entitled "Peculiarities of a Pain in Patients with Ischemic Heart Disease in the Presence of Individual Combines of the Metabolic Syndrome" is awarded Best Article for Vol 12 issue 24
A Study by Sindhu Priya E S et al. entitled "Neuroprotective activity of Pyrazolone Derivatives Against Paraquat-induced Oxidative Stress and Locomotor Impairment in Drosophila melanogaster" is awarded Best Article for Vol 12 issue 23
A Study by Habiba Suhail et al. entitled "Effect of Majoon Murmakki in Dysmenorrhoea (Usre Tams): A Standard Controlled Clinical Study" is awarded Best Article for Vol 12 issue 22
A Study by Ghaffar UB et al. entitled "Correlation between Height and Foot Length in Saudi Population in Majmaah, Saudi Arabia" is awarded Best Article for Vol 12 issue 21
A Study by situs slot and Siti Sarah Binti Maidin entitled "Sleep Well: Mobile Application to Address Sleeping Problems" is awarded Best Article for Vol 12 issue 20
A Study by Avijit Singh et al. situs slot "Comparison of Post Operative Clinical Outcomes Between “Made in India” TTK Chitra Mechanical Heart Valve Versus St Jude Mechanical Heart Valve in Valve Replacement Surgery" is awarded Best Article for Vol 12 issue 19
A Study by Sonali Banerjee and Mary Mathews N. entitled "Exploring Quality of Life and Perceived Experiences Among Couples Undergoing Fertility Treatment in Western India: A Mixed Methodology" is awarded Best Article for Vol 12 issue 18
A Study by Jabbar Desai et al. entitled "Prevalence of Obstructive Airway Disease in Patients with Ischemic Heart Disease and Hypertension" is awarded Best Article for Vol 12 issue 17
A Study by Juna Byun et al. entitled "Study on Difference in Coronavirus-19 Related Anxiety between Face-to-face and Non-face-to-face Classes among University Students in South Korea" is awarded Best Article for Vol 12 issue 16
A Study by Sudha Ramachandra & Vinay Chavan entitled "Enhanced-Hybrid-Age Layered Population Structure (E-Hybrid-ALPS): A Genetic Algorithm with Adaptive Crossover for Molecular Docking Studies of Drug Discovery Process" is awarded Best article for Vol 12 issue 15
A Study by Varsha M. Shindhe et al. entitled "A Study on Effect of Smokeless Tobacco on Pulmonary Function Tests in Class IV Workers of USM-KLE (Universiti Sains Malaysia-Karnataka Lingayat Education Society) International Medical Programme, Belagavi" is awarded Best article of Vol 12 issue 14, July 2020
A study by Amruta Choudhary et al. entitled "Family Planning Knowledge, Attitude and Practice Among Women of Reproductive Age from Rural Area of Central India" is awarded Best Article for special issue "Modern Therapeutics Applications"
A study by Raunak Das entitled "Study of Cardiovascular Dysfunctions in Interstitial Lung Diseas epatients by Correlating the Levels of Serum NT PRO BNP and Microalbuminuria (Biomarkers of Cardiovascular Dysfunction) with Echocardiographic, Bronchoscopic and HighResolution Computed Tomography Findings of These ILD Patients" is awarded Best Article of Vol 12 issue 13 
A Study by Kannamani Ramasamy et al. entitled "COVID-19 Situation at Chennai City – Forecasting for the Better Pandemic Management" is awarded best article for  Vol 12 issue 12
A Study by Muhammet Lutfi SELCUK and Fatma situs slot entitled "Distinction of Gray and White Matter for Some Histological Staining Methods in New Zealand Rabbit's Brain" is awarded best article for  Vol 12 issue 11
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A Study by situs slot et al entitled "Estimation of Reference Interval of Serum Progesterone During Three Trimesters of Normal Pregnancy in a Tertiary Care Hospital of Kolkata" is awarded best article for  Vol 12 issue 09
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A Study by K. Krupashree et. al. entitled "Protective Effects of Picrorhizakurroa Against Fumonisin B1 Induced Hepatotoxicity in Mice" is awarded best article for issue Vol 10 issue 20
A study by Mithun K.P. et al "Larvicidal Activity of Crude Solanum Nigrum Leaf and Berries Extract Against Dengue Vector-Aedesaegypti" is awarded Best Article for Vol 10 issue 14 of IJCRR
A study by Asha Menon "Women in Child Care and Early Education: Truly Nontraditional Work" is awarded Best Article for Vol 10 issue 13
A study by Deep J. M. "Prevalence of Molar-Incisor Hypomineralization in 7-13 Years Old Children of Biratnagar, Nepal: A Cross Sectional Study" is awarded Best Article for Vol 10 issue 11 of IJCRR
A review by Chitra et al to analyse relation between Obesity and Type 2 diabetes is awarded 'Best Article' for Vol 10 issue 10 by IJCRR. 
A study by Karanpreet et al "Pregnancy Induced Hypertension: A Study on Its Multisystem Involvement" is given Best Paper Award for Vol 10 issue 09
Late to bed everyday? You may die early, get depression situs slot
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List of Awardees

A Study by Ese Anibor et al. "Evaluation of Temporomandibular Joint Disorders Among Delta State University Students in Abraka, Nigeria" from Vol 13 issue 16 received Emerging Researcher Award


A Study by Alkhansa Mahmoud et al. entitled "mRNA Expression of Somatostatin Receptors (1-5) in MCF7 and MDA-MB231 Breast Cancer Cells" from Vol 13 issue 06 received Emerging Researcher Award


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International Journal of Current Research and Review (IJCRR) provides platform for researchers to publish and discuss their original research and review work. IJCRR can not be held responsible for views, opinions and written statements of researchers published in this journal

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