Atanda, O.O. and Olopade, T. A .Effect of lemon grass (Cymbopogon citratus (DC.) Stapf.) treatments on Aspergillus flavus (SGS-421) infestation and aflatoxin B1 content of maize grains. International Food Research Journal 20(4): 1933-1939 (2013)

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Studies were conducted to determine the effect of lemon grass (Cymbopogon citratus (DC.) Stapf.) treatments on A. flavus (SGS-421) infestation of maize grains and aflatoxin B1 (AFB1) content of the grains. The experimental design followed a 2×2×5 factorial combination and effect of three factors: form of leaf addition (whole and powdered), form of water suspension (cold and hot), and level of concentration of leaf (0, 8, 10, 12 and 14%, w/w) and water suspension (0, 8, 10, 12 and 14%, w/v); were evaluated by analysis of the maize grains at 5 day interval for 30 days. The growth of the aflatoxigenic fungus and the aflatoxin content of the maize grains were evaluated with palm kernel agar and by thin layer chromatography method respectively. Powdered leaves of lemon grass at 14% (w/w) concentration prevented the growth of the fungus and AFB1 formation in the grains by the 25th day of incubation. This was followed by the whole leaves which reduced the fungal growth to 12.17% and aflatoxin content to 42.62%. Both the cold (8.90%) and hot suspensions (4.15%) had no significant (p > 0.05) effect on the fungal growth after 10 days of incubation. On the contrary, there were significant (p < 0.05) reductions in the aflatoxin content of grains treated with cold (10.24%) and hot suspensions (10.25%) at lowest concentrations of 8%. The findings showed that A. flavus infestation and AFB1 formation in maize grains can be controlled by co-storing powdered leaves of lemon grass with aflatoxin infected maize.

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... For instance, essential oils of basil, anise, clove, cinnamon, lemongrass, lemon, spearmint and oregano have been shown to have inhibitory effects on Fusarium graminearum, F. proliferatum and F. verticillioides that are principal producers of (DON, FB 1 and ZEA in corn and wheat [27]. Cymbopogon citratus (lemongrass) has also been applied for the reduction of Aspergillus species and aflatoxin in maize [28]. Cymbopogon citratus, commonly consumed as tea, is used in the treatment of cold, cough, fever, and stomach upsets [29]. ...
... Five hundred (500) µL of each extract was then filtered with a 0.2 μm Millipore syringe filter and transferred into a Restek 2 mL amber vial (USA) before injecting into the LC-MS/MS system. The formulations were applied at concentrations of 8 and 12% (w/w) in duplicates to millet grains in 250 ml conical flasks and stored at 30 ℃ for four weeks according to the modified method of Atanda and Olopade [28]. A millet sample from the same treatment batch was also subjected to the same storage condition as the treated samples over the same period to serve as a control. ...
... Further statistical analysis using the Least Significant Test (LSD) test to compare mean values among the various treatment formulations for the decontamination of ZEA revealed that lemongrass powder at a concentration of 12% was the most effective treatment against ZEA in millet (p = 0.05). In the study by Atanda and Olopade [28] who applied lemongrass powder for the decontamination of AF in maize, it was revealed that the rate of aflatoxin B 1 (AFB 1 ) decontamination in maize grains was dependent on incubation time and dose of treatment. ...
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Montmorillonite clay due to its abundance and environmental friendliness, has several industrial applications among which is the adsorption of mycotoxins in foods and feeds as binding agents. Fungal species from the genus Fusarium produce zearalenone (ZEA); an oestrogenic compound, which has been implicated in hormonal and reproductive issues for both animal and man. In this study, various nanoformulations from montmorillonite clay and Cymbopogon citratus (lemongrass) extracts were developed for the decontamination of ZEA in millet. The various formulations developed included montmorillonite clay and essential oil of Cymbopogon citratus (Mont-LGEO), montmorillonite clay mixed with C. citratus (lemongrass) powder (Mont-LGP) and montmorillonite clay washed with 1 mM NaCl (Mont-Na). Unmodified montmorillonite clay (Mont) and C. citratus (lemongrass powder) (LGP) served as the negative controls for the treatment. Each of the formulations was exposed to millet grains at concentrations of 8 and 12% and stored for 4 weeks. All the formulations were effective in the decontamination of ZEA in millet after 4 weeks with LGP exposed at 12% recording the highest reduction of 98.3% while the second most effective formulation, Mont-LGP exposed at 12% showed a 66% reduction of ZEA in millet (p = 0.05).
... Other commercial mycotoxin binders employed in feeds include Mycosorb, Formycin, and Anzymit, amongst others [10]. Cymbopogon citratus (lemongrass) powder and extracts are equally effective in decontaminating mycotoxins in crops [11][12][13]. Furthermore, a mixture of montmorillonite clay and lemongrass has been applied for the same purpose in cereal grains. ...
... e millet samples were treated with the nanoformulations (Mont, Mont-Na, Mont-LGEO, and Mont-LGP) at 8% and 12% concentrations in duplicates and put in storage for 4 weeks at a temperature of 30°C in the incubator according to the modified method of Atanda and Olopade [12]. e control (untreated millet sample) from the initial batch was also stored at 30°C just as the treated millet samples. ...
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Thermogravimetric analysis (TGA) was carried out to study the stability of nanoformulations used for the decontamination of mycotoxins. The TGA patterns of the nanoformulations from montmorillonite clay and Cymbopogon citratus (lemongrass) extracts were assessed with temperature ranging from ambient (20°C) to 1000°C. The various nanoformulations studied included unmodified montmorillonite clay (Mont), montmorillonite washed with sodium chloride (Mont-Na), montmorillonite mixed with lemongrass essential oil (Mont-LGEO), and montmorillonite mixed with an equal quantity of lemongrass powder (Mont-LGP). There was no significant difference in the median of the various nanoformulations within 4 weeks at p < 0.05 using the Kruskal-Wallis nonparametric test. For the TGA, the first degradation for montmorillonite clay and the nanoformulations occurred at a temperature between 80 and 101°C and was attributed to the loss of lattice water outside the coordination sphere with a range of 3.5-6.5% weight loss. The second degradation occurred within the temperature of 338 to 344°C, and the third, at a temperature between 640 and 668°C for Mont and the formulations of Mont-Na, Mont-LGEO, and Mont-LGP. There were strong similarities in the degradation patterns of Mont and Mont-Na with the minimum difference being the relatively higher weight loss of the sodium-exchanged cation for Mont-Na at the third degradation step. Hence, the order of stability from the most resistant to the least resistant to degradation is as follows: Mont-LGEO ≥ Mont-Na ≥ Mont ≥ Mont-LGP.
... Clays have been employed in the removal of toxins from foods [8]. Plant extracts have also been employed in the decontamination of mycotoxins from food [9]. However, the combination of clay and plant extracts have not been employed in the decontamination of mycotoxins in foods. ...
... Liquid Chromatography-tandem Mass spectrophotometer (LC-MS/MS) as reported by Sulyok et al. [11] was used to quantify T-2 toxin in the maize sample before treatment with the two types of modified clay (MMT-LGEO and Na-MMT). Each treatment was applied to the maize sample in duplicates at 8% and 12% and stored for 4 weeks at a temperature of 30 o C in the incubator following the modified method of [9]. A control was set up by subjecting the maize sample from the same batch to the same conditions of storage but without any treatment applied. ...
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Montmorillonite clay has a wide range of industrial applications which include the removal of mycotoxins in foods and feed because of its low toxicity both in vitro and in vivo . T-2 toxin is produced mostly by fungal species of Fusarium. Other T-2 producing fungal species are Myrothecium and Stachybotrys . T-2 toxin poses several health hazards such as dystrophy in the brain, heart, kidney and liver as well as ulceration and necrosis of the digestive tract in man and animals. To reduce T-2 toxin in maize, montmorillonite clay modified with lemongrass essential oil (MMT-LGEO) and montmorillonite clay washed with NaCl (Na-MMT) were applied to maize at a concentration of 8% and 12% and kept under storage for one month at 30°C. Untreated maize samples and unmodified montmorillonite clay (MMT) served as controls. The FTIR spectra were recorded for the two treatments and unmodified montmorillonite clay (MMT) used for the removal of T-2 toxin in maize. The FTIR spectra of the two treatments and unmodified clay (MMT) showed the major functional groups as Si-O and -OH. All the treatments reduced the level of T-2 toxin in maize. However, sodium montmorillonite (Na-MMT) and montmorillonite clay modified with lemongrass essential oil (MMT-LGEO) were more efficient than unmodified montmorillonite clay (MMT) in the removal of T-2 toxin in maize.
... The oils of plants such as anise, basil, cinnamon, lemon, clove, lemongrass, spearmint, and oregano have been shown to inhibit fungi that produce DON, FB 1 , and ZEA [27]. Cymbopogon citratus (lemongrass) has been expended in the decontamination of Aspergillus species and limit AF production in maize [28]. Traditionally, lemongrass tea is consumed in South America, Asia, and West Africa, having anti-fever, anti-inflammatory, antiseptic, anti-dyspeptic, and carminative properties [29] with no toxicity reported both in animal and man. ...
... Each of the various treatment formulations (LGEO-MMT, LGP, Na-MMT, MMT, and LGP-MMT) were applied to the maize sample in duplicates at 8% and 12% and stored for 4 weeks at a temperature of 30 • C in the incubator following the modified method of Atanda and Olopade [28]. A control was set up by subjecting the untreated maize sample from the same batch to the same condition of storage at 30 • C, just like the treated maize samples. ...
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Montmorillonite clay has a wide range of applications, one of which includes the binding of mycotoxins in foods and feeds through adsorption. T-2 toxin, produced by some Fusarium, Myrothecium, and Stachybotrys species, causes dystrophy in the brain, heart, and kidney. Various formulations that include lemongrass essential oil-modified montmorillonite clay (LGEO-MMT), lemongrass powder (LGP), montmorillonite clay washed with 1 mM NaCl (Na-MMT), montmorillonite clay (MMT), and lemongrass powder mixed with montmorillonite clay (LGP-MMT) were applied to maize at concentrations of 8% and 12% and stored for a period of one month at 30 °C. Unmodified montmorillonite clay and LGP served as the negative controls alongside untreated maize. Fourier Transform Infrared (FTIR) spectra of the various treatments showed the major functional groups as Si-O and -OH. All treatment formulations were effective in the decontamination of T-2 toxin in maize. Accordingly, it was revealed that the inclusion of Na-MMT in maize at a concentration of 8% was most effective in decontaminating T-2 toxin by 66% in maize followed by LGP-MMT at 12% inclusion level recording a 56% decontamination of T-2 toxin in maize (p = 0.05). Montmorillonite clay can be effectively modified with plant extracts for the decontamination of T-2 toxin.
... Several fungi in grains are pathogenic and are mycotoxin producers. Therefore, various measures have been employed to ensure food sustainability such as fungal decontamination methods using Cymbopogon citratusin maize [14] and mycotoxin decontamination methods in cereal grains using a combination of Montmorillonite clay and Cymbopogon citratus [15,16]. However, the presence of Trametes sp in cereals such as sorghum and millet may not pose a health risk as they have not been reported to produce mycotoxins but rather have potential uses in medicine and the industry. ...
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Trametes species are mushroom fungi with several biotechnological applications. This includes decolourisation of wastewater from olive mill and elimination of endocrine-disrupting hormones. This study reports the presence of two Trametesspecies, namely Trametespolyzona and Trametesvillosa from the phylumBasidiomycota in sorghum and milletvended inSouthwest Nigeria. These fungal isolates were identified culturally and further identified through phylogenetic characterisation. Trametessp occurred in 10% of sorghum samples and 20% of millet samples. The two species were morphologically similar but distantly related phylogenetically. Most fungal species present in cereal crops belong to the division Ascomycota. However, two Trametesspecies belonging to division Basidiomycota are being reported for the first time in cereal crops. Trametessp can be harnessed for their health benefits such as the treatment of cancer and the reduction of viral activity in humans.
... The efficacy of the essential oil in preserving the quality of melon seeds in stores was similar to fungicide (iprodione) treatment. Atanda and Olopade [36] tested potential of C. citratus leaf powder in prevention of Aspergillus flavus infestation in seed treatment of maize and concluded that 14% (w/w) leaf powder prevents of maize grains (25 th day of incubation) and thus disables accumulation of aflatoxin B1 (AFB1) content of the grains. Another study also confirmed in vitro, direct toxic activities of C. citratus essential oil on the mycelial growth and sporulation of Alternaria solani, however when applied onto tomato plants (250, 500, 750, 1000 and 1500 μl/l, 72 h before the fungi inoculation) inhibition of sporulation failed, only fungal growth was prevented and it was in correlation to essential oil concentration [37]. ...
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Plants, particularly medicinal and aromatic species, and plant-derived compounds have been used for centuries in human and veterinary medicine, but nowadays they have increasingly important role in agro-food industry. They present a rich source of bioactive compounds with a wide range of applications that answer to/coup with certain emerging challenges. The most important are growing demands for food safety and concern about human health and environmental pollution that altogether impose the need for more intensive use of plants and their compounds in food industry and agricultural production. This is a reason why, in recent decades, more intensive research has been carried out related to new, bio-rational and specific trends in agro-food industry. Cymbopogon citratus Stapf., lemongrass, is one of medicinal plant species with large application potential in different areas. This review provides an insight into current research and potential applications of C. citratus in food and feed technology, plant protection (as repellent, biofungicide, bioinsecticide, bioherbicide, etc.) and in veterinary medicine purposes. The most comprehensive research on biological activity of lemongrass has been carried out in the field of medicine, entomology and plant protection. However, the information on allelopathic effects and agro-food application is scarce and insufficient, requiring additional research.
... The level of zearalenone in the millet was quantified using the Liquid Chromatography tandem Mass spectrophotometer (LC-MS/MS) as described by [9] before the application of nanoformulations. The formulations were applied to the cereals in duplicates at 8% and 12% and stored at 30 ºC for 4 weeks according to the modified method of [10].A millet sample from the same treatment batch was also subjected to the same storage condition as the treated samples over the same period to serve as a control. The level of mycotoxins in the treated millet with various formulations were analyzed after the storage period of 4 weeks using LC-MS/MS protocol described by [9]. ...
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Montmorillonite clay due to its abundance and environmental friendliness has several industrial applications among which are the adsorption of mycotoxins in foods and feed. The mycotoxin; zearalenone (ZEA) is oestrogenic and has been implicated in hormonal and reproductive issues for both man and animals. Thus, nanoformulations from Montmorillonite clay and Cymbopogoncitratus (lemongrass) extracts were developed for the reduction of the toxin in millet. The various formulations include: Montmorillonite clay to which extract of Cymbopogoncitratus (lemon grass essential oil)has been incorporated (Mont-LGEO), Montmorillonite clay mixed with C. citratus (lemon grass)powder (Mont-LGP) and Montmorillonite clay washed with NaCl (Mont-Na). Pure Montmorillonite clay (Mont) and C. citratus (lemon grass) powder (LGP) served as the controls for the treatment. Each of these compositions were applied to millet samples at a concentration of 8 % and 12% and stored for 4 weeks. XRD patterns for compositions containing Montmorillonite clay revealed major peaks at 2-Theta value of 20.06° representing Montmorillonite and 26.56° and 68.53°representing quartz. The level of ZEA in all treated samples was quantified after 4 weeks using the Liquid Chromatography tandem mass spectrophotometer LC-MS/MS. All the compositions were effective in the decontamination of zearalenone. Furthermore, C. citratus powder was the most effective in the decontamination of zearalenone in the cereal after 4 weeks.
... For example, Agave asperrima and A. striata extracts have been reported to have inhibitory effects on growth of A. flavus and A. parasiticus thus lowering aflatoxin production in affected crops (Sanchez et al., 2005), while oils of Cinnamomum cassia and leaves of Laurus nobilis have been shown to drastically lower aflatoxin production in A. parasiticus CFR 223 without affecting mould growth (Atanda et al., 2007). Lemon grass (whole and powdered leaves, aqueous extracts and volatile compounds) has also been reported to delay fungal growth and inhibit aflatoxin synthesis (Guynot et al., 2003;Atanda and Olopade, 2013). Regardless of the wide application of plant and plant products in the control of fungi and their toxins, literature is scarce on the use of Ashanti pepper (AP) for aflatoxin control. ...
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This study evaluated the influence of co-storing Ashanti pepper (AP), Piper guineense, with maize grains under poor conditions on aflatoxin formation/levels and aflatoxigenic fungal population in the grains. Maize grains were co-stored for 56 days with 1.25%, 2.5% and 5% (w/w) concentrations of either whole AP fruits or powder. Aflatoxin content and population of Aspergillus section Flavi in the grains were analyzed at intervals of 14 days. Aflatoxin formation in the whole and powdered AP–treated grains was significantly (p<0.05) inhibited in a concentration and time dependent manner while reduction in aflatoxigenic fungal population was significantly (p<0.05) more observed in the whole AP–treated grains compared to those co-stored with AP powder. The 2.5% AP powder treatments proved more effective than other powder concentrations against aflatoxin formation (% inhibition = 93.5) at 28 days of storage. However, 5% whole AP treatment was the most effective of all AP treatments against aflatoxin formation (% inhibition = 95.8–99.7) across the 56-day storage period. Co-storing maize grains with whole AP fruits therefore presents a farmer-friendly approach to aflatoxin mitigation during post-harvest storage. [Ezekiel CN, Anokwuru CP, Amos-Tautua BMW, Ejiofor EE, Oriola OR, Obani T, Olajuyigbe OO. Ashanti pepper (Piper guineense) reduces aflatoxin formation in poorly stored maize grain. N Y Sci J 2014;7(9):64-71].
In the present study, in vitro test was carried out to analyze the antifungal potentiality of aqueous extract of five medicinal plants against Fusarium oxysporum f. sp. ciceri by using poisoned food technique at four different concentrations i.e., 10, 25, 50 and 75%. Among all medicinal plant extracts Tinospora cordifolia, Cymbopogon citratus and Moringa oleifera showed the promising antifungal potentiality against F. oxysporum f. sp. ciceri with maximum inhibition i.e., 100% at 75% concentration followed by Zingiber officinale and Trachyspermum ammi, respectively.
The economic burdens and health implications of food spoilage are increasing. Contamination of food sources by fungi, bacteria, yeast, nematodes, insects, and rodents remains a major public health concern. Research has focused on developing safer natural products and innovations to meet consumers' acceptance as alternatives to synthetic food preservatives. Many recent novel preservative techniques and applications of both natural and synthetic origin continue to proliferate in food and chemical industries. In particular, some essential oils of plant origin are potent food preservatives and are thus attractive alternatives to synthetic preservatives. This paper provides an overview of recent advances and future prospects in assessing the efficacy of theuse of Cymbopogon citratus (lemongrass) essential oil in food preservation. The possible mechanisms of action and toxicological profile as well as evidence for or against the use of this essential oil as an alternative to synthetic food preservatives in domestic and industrial applications are discussed.
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A brief account of 50 plant species used by the Kadazandusun communities living around the Crocker Range Sabah, Malaysia is present ed here. It is interesting to note the commonality between plants used to treat specific ailments by the Kadazandusun communities here and other indigenous communities of Borneo as pre viously documented by our own studies. The plants include those used for treating common afflictions such as minor wounds, skin diseases, diarrhea, fever, coughs and malaria. Among the plant species that seem to appear repeatedly in our documentation of medicinal plants in Borneo are Blumea balsamifera for fever, Cassia alata for skin diseases, Centella asiatica for stomachache, Gendarusa vulgaris for general malaise, Nicotiana tabacum as insect repellent, Psidium guajava for diarrhoea, Phyllanthus niruri for malaria, Tinospora crispa for hypertension and Zingiber officinale for rheumatism.
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This review presents the different mycotoxins (aflatoxins, fumonisins and ochratoxin A) produced in agricultural crops in the West African sub-region. The acute and chronic toxic effects of the various mycotoxins are presented. Maize and groundnuts have been found to be excellent substrate for aflatoxin contamination, while fumonisins are widely distributed in maize. Other food products for which mycotoxin contamination has been reported in the sub-region include dried yam chips, tiger nut, melon seeds and stored herbal plants. Mycotoxin contamination is favoured by stress factors during plant growth, late harvesting of crops, high ambient humidity preventing thorough drying, unscientific storage practices and lack of awareness. Control measures include education of the populace on the danger of mycotoxin contaminated diet, early harvesting, rapid drying, sorting, sanitation, use of improved storage structures, smoking, insect control, the use of botanicals and synthetic chemicals as storage protectants, fumigation, biological control, the use of resistant varieties and detoxification of mycotoxin contaminated grains.
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Experiments were carried out to determine the potential of using the powder and essential oil from dried ground leaves of Cymbopogon citratus (lemon grass) to control storage deterioration and aflatoxin contamination of melon seeds (Colocynthis citrullus L.). Four mould species: Aspergillus flavus, A. niger, A. tamarii and Penicillium citrinum were inoculated in the form of conidia suspension (approx. 10 6 conidia per ml) unto shelled melon seeds. The powdered dry leaves and essential oil from lemon grass were mixed with the inoculated seeds at levels ranging from 1-10% (w/w) and 0.1 to 1%v/vt respectively. The ground leaves significantly reduced the extent of deterioration in melon seeds inoculat4ed with different fungi compared to the untreated inoculated seeds. The essential oil at 0.1 and 0.25% (v/w) and ground leaves at 10% (w/w) significantly reduced deterioration and aflatoxin production in shelled melon seeds inoculated with toxigenic A. flavus. At higher dosages (0.5 and 1.0% v/w), the essential completely prevented aflatoxin production. After 6 months in farmers' stores, unshelled melon seeds treated with 0.5% (v/w) of essential oil and 10% (w/w) of powdered leaves of C. citratus had significantly lower proportion of visibly diseased seeds and Aspergillus spp infestation levels and significantly higher seed germination compared to the untreated seeds. The oil content, free fatty acid and peroxide values in seeds protected with essential oil after 6 months did not significantly differ from the values in seed before storage. The efficacy of the essential oil in preserving the quality of melon seeds in stores was statistically at par with that of fungicide (iprodione) treatment.
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Hepatocellular carcinoma (HCC), or liver cancer, is the third leading cause of cancer deaths worldwide, with prevalence 16-32 times higher in developing countries than in developed countries. Aflatoxin, a contaminant produced by the fungi Aspergillus flavus and Aspergillus parasiticus in maize and nuts, is a known human liver carcinogen. We sought to determine the global burden of HCC attributable to aflatoxin exposure. We conducted a quantitative cancer risk assessment, for which we collected global data on food-borne aflatoxin levels, consumption of aflatoxin-contaminated foods, and hepatitis B virus (HBV) prevalence. We calculated the cancer potency of aflatoxin for HBV-postive and HBV-negative individuals, as well as the uncertainty in all variables, to estimate the global burden of aflatoxin-related HCC. Of the 550,000-600,000 new HCC cases worldwide each year, about 25,200-155,000 may be attributable to aflatoxin exposure. Most cases occur in sub-Saharan Africa, Southeast Asia, and China where populations suffer from both high HBV prevalence and largely uncontrolled aflatoxin exposure in food. Aflatoxin may play a causative role in 4.6-28.2% of all global HCC cases.
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The concentration and the chemical composition of the essential oils obtained from different samples of Cymbopogon citratus were evaluated. Among the 12 samples investigated (11 dried leaf samples and fresh plant leaves), seven presented essential oil concentrations within the threshold established by the Brazilian legislation. The moisture content was also determined and the majority of the samples presented humidity contents near 12%. The GC and GC/MS analyses of the essential oils led to identification of 22 compounds, with neral and geranial as the two major components. The total percentage of these two compounds varied within the investigated sample oils from 40.7% to 75.4%. In addition, a considerable variation in the chemical composition of the analyzed samples was observed. The process of grinding the leaves significantly decreased (by up to 68%) the essential oil content, as well as the percentage of myrcene in the oils.
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A scientifically based guide has been developed to evaluate the safety of naturally occurring mixtures, particularly essential oils, for their intended use as flavor ingredients. The approach relies on the complete chemical characterization of the essential oil and the variability of the composition of the oil in the product intended for commerce. Being products of common plant biochemical pathways, the chemically identified constituents are organized according to a limited number of well-established chemical groups called congeneric groups. The safety of the intake of the each congeneric group from consumption of the essential oil is evaluated in the context of data on absorption, metabolism, and toxicology of members of the congeneric group. The intake of the group of unidentified constituents is evaluated in the context of the consumption of the essential oil as a food, a highly conservative toxicologic threshold, and toxicity data on the essential oil or an essential oil of similar chemotaxonomy. The flexibility of the guide is reflected in the fact that high intake of major congeneric groups of low toxicologic concern will be evaluated along with low intake of minor congeneric groups of significant toxicological concern (i.e., higher structural class). The guide also provides a comprehensive evaluation of all congeneric groups and constituents that account for the majority of the composition of the essential oil. The overall objective of the guide is to organize and prioritize the chemical constituents of an essential oil in order that no reasonably possible significant risk associated with the intake of essential oil goes unevaluated. The guide is, however, not intended to be a rigid checklist. The Flavor and Extract Manufacturers Association (FEMA) Expert Panel will continue to evaluate each essential oil on a case by case basis applying their scientific judgment to insure that each natural flavor complex is exhaustively evaluated.
Hepatocellular carcinoma (HCC) is the fifth leading cause of cancer mortality in the world and, in certain parts of Asia and Africa, it accounts for about 70% of cancer deaths. Chronic hepatitis B virus (HBV) infection and dietary exposure to aflatoxin B-1 (AFB(1)) are the two major risk factors in multi factorial aetiology of HCC. Multiple lines of evidence indicate synergistic interaction between these two agents in the development of HCC. Several mechanisms of interaction have been suggested including activation of cytochrome P450s by HBV infection leading to the metabolism of inactive AFB(1) to the mutagenic AFB(1)-8,9-epoxide as well as the generation of reactive oxygen species by HBV and AFB(1) sensitising the cells to AFB(1)-induced p53 249(ser) mutations. The poor survival rate achieved by the current surgical procedures and chemotherapy treatment has motivated a number of scientific investigations to elucidating the molecular events involved in HCC thus providing the scientific rationale for prevention strategies, including primary and chemoprevention approaches. Recent findings have implicated intracellular signalling cascades involving nuclear factor kappa B (NF-kappa B) and nuclear transcription factor erythroid 2p45 (NF-E2)-related factor 2 (Nrf2) as molecular targets of a wide range of chemopreventive agents. The new findings thus raise the intriguing possibility that chemopreventives modulating these molecular targets in the liver might provide a novel therapeutic approach to the development of liver cancer.
The feasibility of using palm kernel agar (PKA) as an alternative culture medium to desiccated coconut agar (DCA), the conventional medium for the recovery of aflatoxigenic fungi from mixed cultures and the detection of aflatoxigenic fungi and direct visual determination of aflatoxins in agricultural commodities was assessed. The medium recovered aflatoxigenic fungi in 48 h from mixed cultures and agricultural commodities in 58 h as compared to 62 h obtained for the two treatments on desiccated coconut medium. Aflatoxigenic fungi were detected in all the agricultural commodities except for onions with maize having the highest value of 13.18% (w/w) followed by melon (10.97), yam flour (10.23) and groundnut (8.52) as against 11.48, 10.0, 6.92 and 8.52% (w/w) obtained for DCA. All aflatoxigenic strains produced a characteristic yellow pigmentation on a pink background and blue or blue green fluorescence of palm kernel agar Under long wave UV light (366nm) as against the white background of DCA, which often interferes with fluorescence with corresponding yield of aflatoxins. This shows that the medium is able to efficiently detect aflatoxin production through direct visual observation of fluorescence. Palm kernel agar (PKA) can therefore be routinely used as an alternative culture medium for screening aflatoxigenic fungi and direct visual determination of aflatoxins in agricultural commodities since it is faster and has a unique pink background for easy identification.
Palm kernel is a cheap natural resource which is abundantly available in the tropics, parts of Asia and South and Central America. A culture medium was developed by incorporating fresh palm kernel extract for the detection of aflatoxigenic fungi. Aflatoxin positive isolates of Aspergilliexhibited a characteristic blue or blue green fluorescence of agar under long wave UV light against a pink background which was confirmed by thin layer chromatography. As compared to conventional desiccated coconut agar, the fluorescent nature of the medium, the intensity and diffusion of the hot water soluble fluorescent compounds of the fungus was unique on this medium. The optimal pH and temperature conditions of aflatoxin production were 7 and 30 ºC respectively. Additives (synthetic and natural) either had no effect or adversely affected the fluorescence of the medium. Aflatoxin detection was possible within 36h in palm kernel broth compared to 40 h in coconut broth. The optimal time of production of fluorescence was 44 h on palm kernel agar compared to 48 h on the conventional medium. Further tests with isolates from different sources showed that yellow pigmentation, fluorescence and aflatoxins were complementary thus obviating the need for UV light. It is thus possible to presumptively identify aflatoxin positive isolates.
The efficacy between analytical and industrial grade antioxidants (butylated hydroxytoluene-BHT, propyl paraben-PP and butylated hydroxyanisole-BHA) at 10 and 20mmolg−1 concentrations on Aspergillus section Flavi populations, other natural competing mycoflora and aflatoxin B1 (AFB1) accumulation at different water activity (aW) levels during 35 days on peanut grains was compared. Assays were carried out on natural peanut grains conditioned at different aW (0.982, 0.955, 0.937). Both grades of BHA–PP mixture M4 (20+20mM) and BHA–PP–BHT mixtures M5 (10+10+10mM), M6 (10+20+10mM), M7 (20+10+10mM) and M8 (20+20+10mM) totally inhibited the growth of Aspergillus section Flavi and peanut mycoflora at all conditions tested. In the same way, BHA–PP mixtures M3 (20+10mM), M4 and BHA–PP–BHT mixtures M5, M6, M7 and M8 completely inhibited the aflatoxin accumulation. The study showed that both antioxidant grades are effective fungal inhibitors to peanut Aspergillus section Flavi populations, natural competing mycoflora and AFB1 accumulation in a wide range of aW during 35 days. The application of the best industrial grade antioxidants mixture tested could be an economic alternative to use in the fungal spoilage control on peanut grains.
The fungitoxicity of crude extracts and essential oils of Achillea millefolium, Cymbopogon citratus, Eucalyptus citriodora and Ageratum conyzoides on the fungus Didymella bryoniae was verified in vitro by means of germination of spores and mycelial growth. In addition, some observations were made using scanning electron microscopy (SEM) to detect possible alterations on the hyphae of Didymella bryoniae. The results revealed that crude extracts of E. citriodora and A. conyzoides were more effective in inhibiting the mycelial growth of D. bryoniae whereas in the germination of spores A. conyzoides and A. millefolium were responsible for most of the inhibition, namely, 52 and 46%, respectively. The essential oils of C. citratus, A. conyzoides and E. citriodora provided 100% inhibition of the mycelial growth and germination of spores of D. bryoniae. SEM observations revealed alterations in the growth pattern of hyphae of D. bryoniae when the essential oil of A. millefolium was present.
Within a storage period of 10 days, samples of maize and cowpea treated with lemon grass powder and essential oil showed no physical deterioration. Off-colour, off-odour and mouldiness however characterized untreated control samples of maize and cowpea. The essential oil of lemon grass also inhibited the growths of moulds like Aspergillus flavus, A. fumigatus, Microphomina phaseoli and Penicillium chrysogenum and bacteria such as Escherichia coli, Pseudomonas aeruginosa, Ps. fluorescens, Bacillus subtilis and Staphylococcus aureus. Phytochemical components like alkaloids, tannins and cardiac glycosides found in the powder are believed to be associated with the preservative and antimicrobial effects of lemon grass.
The effect of processing muthokoi, (a traditional dehulled maize dish in Kenya) on aflatoxin content of naturally contaminated maize was investigated. Dehulling decreased aflatoxin levels by 46.6% (5.5–70%) in maize samples containing 10.7–270 ng/g aflatoxin levels. Soaking muthokoi in 0.2%, 0.5% and 1.0% solutions iati, sodium hypochlorite or ammonium persulphate for 6 or 14 h further decreased aflatoxin contents by 28–72% in maize samples containing 107–363 ng/g aflatoxin levels, and boiling muthokoi at 98 °C for 150 min in 0.2–1.0% w/v iati decreased aflatoxin contents by 80–93% in samples having 101 ng/g aflatoxin contamination. Findings imply that exposure to acute aflatoxin levels in maize is minimised during processing and preparation of muthokoi.
The chemical method for confirmation of the identity of aflatoxin by derivative formation directly on the TLC plate was studied collaboratively by 8 participants. The results show that aflatoxin B-1 was confirmed in 17 of 17 sample extracts representing 15 mu-g aflatoxin B-1/kg peanut butter, in 13 of 16 extracts representing 5 mu-g/kg, and in none of the 7 aflatoxin-free extracts. Collaborators commented that the method was easily performed and gave good results. The method has been adopted as official first action.
An infusion (abafado) prepared from leaves of lemongrass (Cymbopogon citratus Stapf) administered orally to adult rats for 2 months, in doses up to 20 times larger than the estimated corresponding human dosage, did not induce any effect which could be taken as evidence of toxicity. An absence of effects was also noted in male and female rats and in their offspring when the abafado was administered prior to mating or during pregnancy. These data strongly suggest that lemongrass, as used in Brazilian folk medicine, has no toxic properties.
Aflatoxin production and degradation were examined in three isolates of A. parasiticus. Maximum yields were present after incubation for 14 days and these declined gradually as the culture aged. Young mycelia (4 days old) synthesized the greatest amounts of aflatoxin, but aging mycelia (14 days old) were mainly responsible for degradation. Addition of cycloheximide to young cultures and removal of mycelia from aging cultures both prevented further aflatoxin degradation. Intramycelial substances released from fragmented or homogenized mycelium were capable of degrading aflatoxins, and their concentration increased as the mycelium aged. When 14C-labelled aflatoxin was added to a 2-day-old culture and further incubated, 75% of the radioactivity at 12 days was intramycelial, but at 20 days, most radioactivity was in the filtrate.
To investigate the volatile fractions of 16 essential oils for activity against the more common fungi causing spoilage of bakery products, Eurotium amstelodami, E. herbariorum, E. repens, E. rubrum, Aspergillus flavus, A. niger and Penicillium corylophilum. The study applied 50 microl of pure essential oils in a sterilized filter paper, were carried out at pH 6 and at different water activity levels (0.80-0.90). First, a wheat flour based agar medium was used, where cinnamon leaf, clove, bay, lemongrass and thyme essential oils where found to totally inhibit all microorganisms tested. These five essential oils were then tested in sponge cake analogues, but the antifungal activity detected was much more limited. Five essential oils showed potential antifungal capacity against all species tested, over a wide range of water availability. Their activity, however, seems to be substrate-dependent. More research is needed to make them work in real bakery products, as in the preliminary study limited effectiveness was found. The potential of the cinnamon leaf, clove, bay, lemongrass and thyme essential oils against species belonging to Eurotium, Aspergillus and Penicillium genus has been demonstrated.
To develop a natural fungicide against aflatoxigenic fungi, to protect stored rice, using the essential oil of lemongrass. Aspergillus flavus Link. was isolated from stored rice and identified as an aflatoxigenic strain. Lemongrass oil was tested against A. flavus and the test oil was fungistatic and fungicidal against the test pathogen at 0.6 and 1.0 mg ml(-1), respectively. Aflatoxin production was completely inhibited at 0.1 mg ml(-1). The results obtained from the thin layer chromatographic bioassay and gas chromatography indicated citral a and b as the fungicidal constituents in lemongrass oil. During the fumigant toxicity assay of lemongrass oil, the sporulation and the mycelial growth of the test pathogen were inhibited at the concentrations of 2.80 and 3.46 mg ml(-1), respectively. Lemongrass oil could be used to manage aflatoxin formation and fungal growth of A. flavus in stored rice. Currently, fungicides are not used to control fungal pests or mycotoxin production on stored rice. Rice treated with the essential oil of lemongrass could be used to manage fungal pests as well as the insect pests in stored rice. The essential oil is chemically safe and acceptable to consumers, as synthetic chemical fungicides can cause adverse health effects to consumers.
Aflatoxins are a family of fungal toxins that are carcinogenic to man and cause immunosuppression, cancer and growth reduction in animals. We conducted a cross-sectional study among 480 children (age 9 months to 5 years) across 4 agro-ecological zones (SS, NGS, SGS and CS) in Benin and Togo to identify the effect of aflatoxin exposure on child growth and assess the pattern of exposure. Prior reports on this study [Gong, Y.Y.,Cardwell, K., Hounsa, A., Egal, S., Turner, Hall, A.J., Wild, C.P., 2002. Dietary aflatoxin exposure and impaired growth in young children from Benin and Togo: cross sectional study. British Medical Journal 325, 20-21, Gong, Y.Y., Egal, S., Hounsa, A., Turner, P.C., Hall, A.J., Cardwell, K., Wild, C.P., 2003. Determinants of aflatoxin exposure in young children from Benin and Togo, West Africa: the critical role of weaning and weaning foods. International Journal of Epidemiology, 32, 556-562] showed that aflatoxin exposure among these children is widespread (99%) and that growth faltering is associated with high blood aflatoxin-albumin adducts (AF-alb adducts), a measure of recent past exposure. The present report demonstrates that consumption of maize is an important source of aflatoxin exposure for the survey population. Higher AF-alb adducts were correlated with higher A. flavus (CFU) infestation of maize (p=0.006), higher aflatoxin contamination (ppb) of maize (p<0.0001) and higher consumption frequencies of maize (p=0.053). The likelihood of aflatoxin exposure from maize was particularly high in agro-ecological zones where the frequency of maize consumption (SGS and CS), the presence of aflatoxin in maize (SGS) or the presence of A. flavus on maize (NGS and SGS) was relatively high. Socio-economic background did not affect the presence of A. flavus and aflatoxin in maize, but better maternal education was associated with lower frequencies of maize consumption among children from the northernmost agro-ecological zone (SS) (p=0.001). The impact of groundnut consumption on aflatoxin exposure was limited in this population. High AF-alb adduct levels were correlated with high prevalence of A. flavus and aflatoxin in groundnut, but significance was weak after adjustment for weaning status, agro-ecological zone and maternal socio-economic status (resp. p=0.091 and p=0.083). Ingestion of A. flavus and aflatoxin was high in certain agro-ecological zones (SS and SGS) and among the higher socio-economic strata due to higher frequencies of groundnut consumption. Contamination of groundnuts was similar across socio-economic and agro-ecological boundaries. In conclusion, dietary exposure to aflatoxin from groundnut was less than from maize in young children from Benin and Togo. Intervention strategies that aim to reduce dietary exposure in this population need to focus on maize consumption in particular, but they should not ignore consumption of groundnuts.
The inhibitory effect of cowdung fumes, Captan, leaf powder of Withania somnifera, Hyptis suaveolens, Eucalyptus citriodora, peel powder of Citrus sinensis, Citrus medica and Punica granatum, neem cake and pongamia cake and spore suspension of Trichoderma harzianum and Aspergillus niger on aflatoxin B(1) production by toxigenic strain of Aspergillus flavus isolated from soybean seeds was investigated. Soybean seed was treated with different natural products and fungicide captan and was inoculated with toxigenic strain of A. flavus and incubated for different periods. The results showed that all the treatments were effective in controlling aflatoxin B(1) production. Captan, neem cake, spore suspension of T. harzianum, A. niger and combination of both reduced the level of aflatoxin B(1) to a great extent. Leaf powder of W. somnifera, H. suaveolens, peel powder of C. sinensis, C. medica and pongamia cake also controlled the aflatoxin B(1) production. All the natural product treatments applied were significantly effective in inhibiting aflatoxin B(1) production on soybean seeds by A. flavus. These natural plant products may successfully replace chemical fungicides and provide an alternative method to protect soybean and other agricultural commodities from aflatoxin B(1) production by A. flavus.
We recently reported that chlorophyll (Chl) strongly inhibits aflatoxin B(1) preneoplasia biomarkers in rats when administered by co-gavage (Simonich et al., 2007. Natural chlorophyll inhibits aflatoxin B1-induced multi-organ carcinogenesis in the rat. Carcinogenesis 28, 1294-1302.). The present study extends this by examining the effects of dietary Chl on tumor development, using rainbow trout to explore ubiquity of mechanism. Duplicate groups of 140 trout were fed diet containing 224 ppm dibenzo[a,l]pyrene (DBP) alone, or with 1000-6000 ppm Chl, for 4 weeks. DBP induced high tumor incidences in liver (51%) and stomach (56%), whereas Chl co-fed at 2000, 4000 or 6000 ppm reduced incidences in stomach (to 29%, 23% and 19%, resp., P<0.005) and liver (to 21%, 28% and 26%, resp., P<0.0005). Chlorophyllin (CHL) at 2000 ppm gave similar protection. Chl complexed with DBP in vitro (2Chl:DBP, K(d1)=4.44+/-0.46 microM, K(d2)=3.30+/-0.18 microM), as did CHL (K(d1)=1.38+/-0.32 microM, K(d2)=1.17+/-0.05 microM), possibly explaining their ability to inhibit DBP uptake into the liver by 61-63% (P<0.001). This is the first demonstration that dietary Chl can reduce tumorigenesis in any whole animal model, and that it may do so by a simple, species-independent mechanism.
Caffeic acid (3,4-dihydroxycinnamic acid, 12 mM) added to a fat-based growth medium reduces >95% of aflatoxin production by Aspergillus flavus NRRL 3357, without affecting fungal growth. Microarray analysis of caffeic acid-treated A. flavus indicated expression of almost all genes in the aflatoxin biosynthetic cluster were down-regulated, ranging from a log2 ratio of caffeic acid treated and untreated of -1.12 (medium) to -3.13 (high). The only exceptions were genes norB and the aflatoxin pathway regulator-gene, aflJ, which showed low expression levels in both treated and control fungi. The secondary metabolism regulator-gene, laeA, also showed little change in expression levels between the fungal cohorts. Alternatively, expression of genes in metabolic pathways (i.e., amino acid biosynthesis, metabolism of aromatic compounds, etc.) increased (log2 ratio >1.5). The most notable up-regulation of A. flavus expression occurred in four genes that are orthologs of the Saccharomyces cerevisiae AHP1 family of genes. These genes encode alkyl hydroperoxide reductases that detoxify organic peroxides. These increases ranged from a log2 ratio of 1.08 to 2.65 (moderate to high), according to real-time quantitative reverse transcription-PCR (qRT-PCR) assays. Based on responses of S. cerevisiae gene deletion mutants involved in oxidative stress response, caffeic, chlorogenic, gallic and ascorbic acids were potent antioxidants under oxidative stress induced by organic peroxides, tert-butyl and cumene hydroperoxides. Differential hypersensitivity to these peroxides and hydrogen peroxide occurred among different mutants in addition to their ability to recover with different antioxidants. These findings suggest antioxidants may trigger induction of genes encoding alkyl hydroperoxide reductases in A. flavus. The possibilities that induction of these genes protects the fungus from oxidizing agents (e.g., lipoperoxides, reactive oxygen species, etc.) produced during host-plant infection and this detoxification attenuates upstream signals triggering aflatoxigenesis are discussed.
Maize samples were collected during a survey in three agro-ecological zones in Nigeria to determine the distribution and aflatoxin-producing potential of members of Aspergillus section Flavi. The three agro-ecological zones were, Derived Savannah (DS) and Southern Guinea Savannah (SGS) in the humid south and North Guinea Savannah (NGS) in the drier north. Across agro-ecological zones, Aspergillus was the most predominant fungal genera identified followed by Fusarium with mean incidences of 70 and 24%, respectively. Among Aspergillus, A. flavus was the most predominant and L-strains constituted >90% of the species identified, while the frequency of the unnamed taxon S(BG) was <3%. The incidence of atoxigenic strains of A. flavus was higher in all the districts surveyed except in the Ogbomosho and Mokwa districts in DS and SGS zones, respectively, where frequency of toxigenic strains were significantly (P<0.05) higher than that of atoxigenic strains. The highest and lowest incidence of aflatoxin positive samples was recorded in the SGS (72%) and NGS (20%), respectively. Aflatoxin contamination in grain also followed a similar trend and the highest mean levels of B-aflatoxins were detected in maize samples obtained from Bida (612 ng g(-1)) and Mokwa (169 ng g(-1)) districts, respectively, in the SGS. Similarly, the highest concentrations of G-aflatoxins were detected in samples from Akwanga district in the SGS with a mean of 193 and 60 ng g(-1), respectively. When agro-ecological zones were compared, B-aflatoxins were significantly (P<0.05) higher in SGS than in NGS, and intermediate in maize samples from the DS agro-ecological zone.
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