New insights into the persistent effect of transient cinnamaldehyde vapor treatment on the growth and aflatoxin synthesis of Aspergillus flavus.

The antimicrobial effects of continuous treatment with essential oils (EOs) in both liquid and gaseous phases have been intensively studied. Due to their rapid volatility, the effects of EOs on microorganisms after transient treatment are also worth exploring. In this work, the persistent effects of cinnamaldehyde (CA) vapor on Aspergillus flavus were detected by a series of biochemical analyses. Transcriptome analysis was also conducted to study the gene expression changes between recovered and normal A. flavus. When CA vapor was removed, biochemical analyses showed that the oxidative stress induced by the antimicrobial atmosphere was alleviated, and almost all the damaged functions were restored apart from mitochondrial function. Remarkably, the suppressed aflatoxin production intensified, which was confirmed by the up-regulation of most genes in the aflatoxin synthetic gene cluster, the velvet-related gene FluG and the aflatoxin precursor acetyl-CoA. Transcriptomic analysis also demonstrated significant changes in secondary metabolism, energy metabolism, oxidative stress, and amino acid metabolism in the recovery group. Taken together, these findings provide new insights into the mechanisms underlying the response of A. flavus to CA vapor treatment and will guide the rational application of EOs.

Protective Effect of SeMet on Liver Injury Induced by Ochratoxin A in Rabbits.

Ochratoxin A (OTA) is second only to aflatoxin in toxicity among mycotoxins. Recent studies have shown that selenomethionine (SeMet) has a protective effect on mycotoxin-induced toxicity. The purpose of this study was to investigate the protective effect and mechanism of SeMet on OTA-induced liver injury in rabbits. Sixty 35-day-old rabbits with similar body weight were randomly divided into five groups: control group, OTA group (0.2 mg/kg OTA), OTA + 0.2 mg/kg SeMet group, OTA + 0.4 mg/kg SeMet group and OTA + 0.6 mg/kg SeMet group. Rabbits were fed different doses of the SeMet diet for 21 d, and OTA was administered for one week from day 15 (the control group was provided the same dose of NaHCO3 solution). The results showed that 0.4 mg/kg SeMet could significantly improve the liver injury induced by OTA poisoning. SeMet supplementation can improve the changes in physiological blood indexes caused by OTA poisoning in rabbits and alleviate pathological damage to the rabbit liver. SeMet also increased the activities of SOD, GSH-Px and T-AOC and significantly decreased the contents of ROS, MDA, IL-1ß, IL-6 and TNF-α, effectively alleviating the oxidative stress and inflammatory response caused by OTA poisoning. In addition, OTA poisoning inhibits Nrf2 and HO-1 levels, ultimately leading to peroxide reaction, while SeMet activates the Nrf2 signaling pathway and enhances the expression of the HO-1 downstream Nrf2 gene. These results suggest that Se protects the liver from OTA-induced hepatotoxicity by regulating Nrf2/HO-1 expression.

Spent Coffee Grounds Valorization as Bioactive Phenolic Source Acquired Antifungal, Anti-Mycotoxigenic, and Anti-Cytotoxic Activities.

Spent coffee grounds (SCGs), which constitute 75% of original coffee beans, represent an integral part of sustainability. Contamination by toxigenic fungi and their mycotoxins is a hazard that threatens food production. This investigation aimed to examine SCGs extract as antimycotic and anti-ochratoxigenic material. The SCGs were extracted in an eco-friendly way using isopropanol. Bioactive molecules of the extract were determined using the UPLC apparatus. The cytotoxicity on liver cancer cells (Hep-G2) showed moderate activity with selectivity compared with human healthy oral epithelial (OEC) cell lines but still lower than the positive control (Cisplatin). The antibacterial properties were examined against pathogenic strains, and the antifungal was examined against toxigenic fungi using two diffusion assays. Extract potency was investigated by two simulated models, a liquid medium and a food model. The results of the extract showed 15 phenolic acids and 8 flavonoids. Rosmarinic and syringic acids were the most abundant phenolic acids, while apigenin-7-glucoside, naringin, epicatechin, and catechin were the predominant flavonoids in the SCGs extract. The results reflected the degradation efficiency of the extract against the growth of Aspergillus strains. The SCGs recorded detoxification in liquid media for aflatoxins (AFs) and ochratoxin A (OCA). The incubation time of the extract within dough spiked with OCA was affected up to 2 h, where cooking was not affected. Therefore, SCGs in food products could be applied to reduce the mycotoxin contamination of raw materials to the acceptable regulated limits.

Structure Elucidation and Toxicity Analysis of the Byproducts Formed after Biodegradation of Aflatoxins B1 and B2 Using Extracts of Mentha arvensis.

The aqueous extracts of leaves and shoots of Mentha arvensis were checked for their potential to biodegrade aflatoxin B1 and B2 (AFB1; 100 µg/L and AFB2; 50 µg/L) through in vitro assays. Overall, the results showed that leaf extract degrades aflatoxins more efficiently than the shoot extract. First, the pH, temperature and incubation time were optimized for maximum degradation by observing this activity at different temperatures between 25 and 60 °C, pH between 2 and 10 and incubation time from 3 to 72 h. In general, an increase in all these parameters significantly increased the percentage of biodegradation. In vitro trials on mature maize stock were performed under optimized conditions, i.e., pH 8, temperature 30 °C and an incubation period of 72 h. The leaf extract resulted in 75% and 80% biodegradation of AFB1 and AFB2, respectively. Whereas the shoot extract degraded both toxins up to 40-48%. The structural elucidation of degraded toxin products by LCMS/MS analysis showed seven degraded products of AFB1 and three of AFB2. MS/MS spectra showed that most of the products were formed by the loss of the methoxy group from the side chain of the benzene ring, the removal of the double bond in the terminal furan ring and the modification of the lactone group, indicating less toxicity compared to the parent compounds. The degraded products showed low toxicity against brine shrimps, confirming that M. arvensis leaf extract has significant potential to biodegrade aflatoxins.

Effects of licorice extract, probiotic, toxin binder and poultry litter biochar on performance, immune function, blood indices and liver histopathology of broilers exposed to aflatoxin-B1.

Probiotics, toxin binders, and plant extracts improve health and immunity of broiler chickens exposed to aflatoxin. The effects of licorice extract (LE), Protexin probiotic, toxin binder (Agrabound), and poultry litter biochar (PLB) in experimental aflatoxicosis were evaluated. In a completely randomized design, 504 broiler chickens were allotted to 7 treatments and 6 replicates with 12 broiler chickens in each. The experimental groups were as follows: T1) basal diet (B) without any feed additive or aflatoxin B1 (AFB1); T2) B + 0.5 mg AFB1/kg; T3) T2 + 3 g LE/kg; T4) T2 + 6 g LE/kg; T5) T2 + 0.5 g Protexin/kg; T6) T2 + 1 g toxin binder/kg, and T7) T2 + 5 g/kg PLB. Broiler chickens fed AFB diet (T2) had lower body weight gain at the end of grower period and higher feed conversion ratio at the end of the finisher period, whereas inclusion of LE, probiotic, toxin binder, or PLB restores body weight of broiler chickens to that of the control group. Aflatoxicosis decreased total protein, TG, albumin, Ca, and P concentrations and greater uric acid concentration in broiler chickens as compared with the control group (P < 0.05). As compared with the T2 group, inclusion of 3 mg LE/kg increased serum total protein; inclusion of 3 mg LE/kg, probiotic, and toxin binder increased TG; inclusion of 3 and 6 mg LE/kg, probiotic, and PLB increased serum albumin; and the whole additive decreased serum uric acid of broiler chickens comparing with the control group. Lymphocyte percentage, avian influenza antibody titer, thymus relative weight, and immune response to phytohemagglutinin were decreased in the T2 group, whereas heterophil percentage and heterophil-to-lymphocyte ratio were increased (P < 0.05). Aflatoxicosis increased breast meat malondialdehyde concentration, liver enzymes activities, and number of fat vacuoles (P < 0.05). As compared with the T2 group, all of the additives lowered alkaline phosphatase, aspartate aminotransferase, and alanine transaminase activities, breast meat malondialdehyde concentration, and liver pathological damages (P < 0.05). It can be concluded that all of the additives are capable to decrease the negative impact of AFB1 on broiler chickens' performance, blood indices, and immunity.

Calcination Improves the In Vivo Efficacy of a Montmorillonite Clay to Bind Aflatoxin G1 in Broiler Chickens: A Toxicokinetic Approach.

The goal of this study was to investigate the toxicokinetic characteristics of aflatoxin G1 (AFG1) in broiler chickens and the effect of calcination of a Tunisian montmorillonite clay on the in vivo absorption of AFG1. In this study, broiler chickens were randomly distributed into four groups of 10 animals. Group 1 was administered AFG1 (2 mg/kg body weight (BW)) by single intravenous injection (IV), group 2 received an intra-crop bolus (PO) of AFG1 without any clay, group 3 was dosed AFG1 PO together with an oral bolus of purified clay (CP), and group 4 received AFG1 PO with an oral bolus of calcined clay. A significant difference in the area under the curve (AUC0-t) was observed for group 4 (6.78 ± 4.24 h*ng/mL) in comparison with group 2 (12.83 ± 4.19 h*ng/mL). A significant reduction of the oral bioavailability of AFG1 was observed for group 4 (7.61 ± 4.76%) compared with group 2 (14.40 ± 4.70%), while no significant effect was observed of CP. In this experiment, no phase I nor phase II metabolites of AFG1 were observed. These findings confirm that calcination of the purified montmorillonite clay enhances the adsorption of AFG1 in the gastrointestinal tract after oral administration, thereby reducing its bioavailability, thus reducing its toxic effects.

Determination of Antioxidant and Antifungal Activities in Cookies Fortified with Solar Dried Prickly Pear Peels Powder.

BACKGROUND AND OBJECTIVE: The fortification of bakery products by new materials that attain various goals is considered a challenging that finally gains useful health amelioration. This study was planned to assess the effect of incorporation of solar dried prickly pear peels powder in qaraqeesh (Egyptian cookies) with respect to increase shelf life, sensory palatability and nutritional value. Prickly pear cactus (Opuntia ficus-indica) beside distributed in arid and semiarid regions proved to have phytochemical compounds with high antioxidants capacity. MATERIALS AND METHODS: Fungi colonies were isolated from prickly pear peels. Three levels (1, 3 and 5%) of dried peels powder were added to wheat flour along with other ingredients to make cookies samples. Mycological analysis was assessed in yeast with the three concentrations of peels powder as well as the fresh peels and negative control. The total phenolics, flavonoids, tannins, anthocyanins and carotenoids as well as the antioxidant activity were evaluated in fresh and dried cactus peels. RESULTS: Findings showed that the prickly pear peels powder (PPPP) antioxidant activity was not much affected by the solar drying conditions. The effect of different extracting solvents at different polarties and pH on the phenolic and flavonoids contents of PPPP was studied. Aflatoxins production by aflatoxignicity A. flavus (ATCC 28542) was inhibited by adding different concentrations of PPPP to cookies. Sensory evaluation of fortified cookies was done. All the evaluated characteristics of cookies were given nearly the same values for all levels of dried peels powder. CONCLUSION: Addition of 5% dried cactus peel had lower overall quality and color than the control. Adding 3% of PPPP to cookies (qaraqeesh) showed the highest sensory score. Dried cactus peels may improve quality, nutritional value and shelf life of cookies.

Assessment of chemically characterised Myristica fragrans essential oil against fungi contaminating stored scented rice and its mode of action as novel aflatoxin inhibitor.

The study reports chemically characterised Myristica fragrans essential oil (MFEO) as plant based food preservative against fungal and aflatoxin B1 (AFB1) contamination of scented rice varieties. The chemical profile of MFEO revealed elemicin (27.08%), myristicine (21.29%) and thujanol (18.55%) as major components. The minimum inhibitory and minimum aflatoxin inhibitory concentrations of MFEO were 2.75 and 1.5 mg/ml, respectively. The MFEO was efficacious against a broad spectrum of food deteriorating fungi. MFEO caused decrease in ergosterol content of fungal plasma membrane and enhanced leakage of cellular ions, depicting plasma membrane as the site of action. The MFEO caused reduction in cellular methylglyoxal content, the aflatoxin inducer. This is the first report on MFEO as aflatoxin suppressor. The essential oil may be recommended as plant based food preservative after large scale trials and reduction in methylglyoxal suggests its application for development of aflatoxin resistant varieties through green transgenics.

Antifungal and antimycotoxigenic metabolites from native plants of northwest Argentina: isolation, identification and potential for control of Aspergillus species.

Extracts from aerial parts of Prosopis ruscifolia, Bidens pilosa, Cercidium praecox and Phoradendron liga were assayed against toxigenic Aspergillus species. They were obtained by sequential extraction of the aerial parts with hexane (fHex), dichloromethane (fDCM), ethyl acetate (fEtOAc) and methanol (fMeOH). The fMeOH from P. ruscifolia showed the highest antifungal spectrum (MIC = 750-1500 µg mL-1; MID = 50-200 µg; DI = 1.7-3.0 mm). Indolizidine alkaloids (juliflorine and juliprosine) and tryptamine were identified with strong (MIC = 188 µg mL-1) and moderate antifungal activities (MIC = 750 µg mL-1), respectively, towards A. parasiticus and A. flavus. The fMeOH, the indolizidine alkaloids and tryptamine synergized the fungitoxic effect of potassium sorbate and propiconazole. They completely suppressed the biosynthesis of aflatoxins at concentrations of 47, 94 and 375 µg mL-1, respectively. Our results indicate that fMeOH and its identified alkaloids are promisory additives of commercial antifungals and are antiaflatoxigenic agents at concentrations below of those required for complete suppression of fungal growth.

Identification and control of specific aflatoxin-producing fungi in stored maize seeds in awka using azadirachta indica (neem) and garcinia kola seeds.

Four fungal isolates were identified in this study of which three were Aspergillus species with Aspergillus flavus having the highest frequency followed by A. parasiticus. The result of high frequency of Aspergillus flavus and Aspergillus parasiticus in the Zea mays sample revealed production of aflatoxins. Maize sample in Awka was found to contain aflatoxin B1 (9.60ppb) and B2 (13.3ppb). Inhibition of A. flavus and A. parasiticus with Azadirachta indica and Garcinia kola seed extracts showed that the test plant extracts were effective for reducing mycelial growth on the test organism. Methanolic extract of G. kola showed antifungal inhibitory activity on the test organisms and the highest at 10% concentration. With ethanol extracts of G. kola, the antifungal activity was effective i.e. for inhibition of A. flavus and A. parasiticus, with A. parasiticus having the higher percentage inhibition at 10%. Inhibiting growth of Aspergillus flavus and Aspergillus parasiticus using methanolic and ethanolic extracts of neem seeds was effective in the inhibition of the test organism at 10%. The methanolic and ethanolic extracts of combined Garcinia kola and neem seeds revealed effective inhibition of A. flavus and A. parasiticus with ethanolic extracts of the combined test plants exerting the highest inhibition against A. flavus (80.43±3.62). The extracts from this plant show the ability to suppress growth of toxigenic A. flavus and A. parasiticus. Phytochemical analysis showed that the methanolic and ethanolic extracts of G. kola and neem seeds showed the presence of secondary metabolites and this may be a reason for the inhibitory activity on A. flavus and A. parasiticus. Results from this study will be important in planning a management strategy against aflatoxin-producing fungi and other fungi associated with spoilage of stored food products.

Aspergillus flavus as a Model System to Test the Biological Activity of Botanicals: An Example on Citrullus colocynthis L. Schrad. Organic Extracts.

Citrullus colocynthis L. Schrader is an annual plant belonging to the Cucurbitaceae family, widely distributed in the desert areas of the Mediterranean basin. Many pharmacological properties (anti-inflammatory, anti-diabetic, analgesic, anti-epileptic) are ascribed to different organs of this plant; extracts and derivatives of C. colocynthis are used in folk Berber medicine for the treatment of numerous diseases-such as rheumatism arthritis, hypertension bronchitis, mastitis, and even cancer. Clinical studies aimed at confirming the chemical and biological bases of pharmacological activity assigned to many plant/herb extracts used in folk medicine often rely on results obtained from laboratory preliminary tests. We investigated the biological activity of some C. colocynthis stem, leaf, and root extracts on the mycotoxigenic and phytopathogenic fungus Aspergillus flavus, testing a possible correlation between the inhibitory effect on aflatoxin biosynthesis, the phytochemical composition of extracts, and their in vitro antioxidant capacities.

Metal Ions in Activated Carbon Improve the Detection Efficiency of Aflatoxin-Producing Fungi.

Aflatoxins (AF), produced by several Aspergillus species, are visible under ultraviolet light if present in high amounts. AF detection can be improved by adding activated carbon, which enhances the observation efficiency of weakly AF-producing fungi. However, commercial activated carbon products differ in their characteristics, making it necessary to investigate which characteristics affect method reproducibility. Herein, the addition of 10 activated carbon products resulted in different AF production rates in each case. The differences in the production of aflatoxin G1 (AFG1) were roughly correlated to the observation efficiency in the plate culture. Trace element analysis showed that the concentrations of several metal ions differed by factors of >100, and the carbons that most effectively increased AFG1 production contained higher amounts of metal ions. Adding 5 mg L-1 Fe or Mg ions increased AFG1 production even without activated carbon. Furthermore, co-addition of both ions increased AFG1 production stably with the addition of carbon. When varying the concentration of additives, only AFG1 production increased in a concentration-dependent manner, while the production of all the other AFs decreased or remained unchanged. These findings suggest that a key factor influencing AF production is the concentration of several metal ions in activated carbon and that increasing AFG1 production improves AF detectability.

Improvement of the Culture Medium for the Dichlorvos-Ammonia (DV-AM) Method to Selectively Detect Aflatoxigenic Fungi in Soil.

The dichlorvos-ammonia (DV-AM) method is a simple but sensitive visual method for detecting aflatoxigenic fungi. Here we sought to develop a selective medium that is appropriate for the growth of aflatoxigenic fungi among soil mycoflora. We examined the effects of different concentrations of carbon sources (sucrose and glucose) and detergents (deoxycholate (DOC), Triton X-100, and Tween 80) on microorganisms in soils, using agar medium supplemented with chloramphenicol. The results demonstrated that 5⁻10% sucrose concentrations and 0.1⁻0.15% DOC concentrations were appropriate for the selective detection of aflatoxigenic fungi in soil. We also identified the optimal constituents of the medium on which the normal rapid growth of Rhizopus sp. was completely inhibited. By using the new medium along with the DV-AM method, we succeeded in the isolation of aflatoxigenic fungi from non-agricultural fields in Fukui city, Japan. The fungi were identified as Aspergillus nomius based on their calmodulin gene sequences. These results indicate that the new medium will be useful in practice for the detection of aflatoxigenic fungi in soil samples including those from non-agricultural environments.

Chemically characterized Mentha cardiaca L. essential oil as plant based preservative in view of efficacy against biodeteriorating fungi of dry fruits, aflatoxin secretion, lipid peroxidation and safety profile assessment.

The study reports Mentha cardiaca essential oil (EO) as plant based preservative against fungal and aflatoxin contamination of stored dry fruits. Mycoflora analysis of the dry fruits revealed Aspergillus favus LHP-PV-1 as the most aflatoxigenic isolate with highest Aflatoxin B1 content. M. cardiaca EO showed broad fungitoxic spectrum inhibiting the tested moulds contaminating dry fruits. It's minimum inhibitory concentration (MIC), minimum aflatoxin inhibitory concentration (MAIC) and minimum fungicidal concentration (MFC) against A. favus LHP-PV-1 were recorded to be 1.25, 1.0 and 2.25 µL/mL respectively. The EO caused decrease in ergosterol content and enhanced leakage of Ca2+, K+ and Mg2+ ions from treated fungal cells, depicting fungal plasma membrane as the site of antifungal action. The EO showed promising DPPH free radical scavenging activity (IC50 value:15.89 µL/mL) and favourable safety profile with LD50 value (7133.70 mg/kg body wt.) when estimated through acute oral toxicity on mice. Carvone (61.62%) was recorded as the major component of the oil during chemical characterisation through GC-MS. Based on strong antifungal, antiaflatoxigenic and antioxidant potential, the chemically characterised M. cardiaca EO may be recommended as safe plant based preservative and shelf life enhancer of food items. This is the first report on antifungal and antiaflatoxigenic activity of M. cardiaca EO.

Positive and negative aspects of green coffee consumption - antioxidant activity versus mycotoxins.

BACKGROUND: The quality of coffee depends not only on the contents of healthy compounds but also on its contamination with microorganisms that can produce mycotoxins during development, harvesting, preparation, transport and storage. RESULTS: The antioxidant activity of green coffee brews measured in this study by ABTS, DPPH and Folin-Ciocalteu assays showed that coffee extracts from Robusta beans possessed higher activity in all assays than extracts from Arabica beans. The occurrence of ochratoxin A and aflatoxins (B1, B2, G1 and G2) in green coffee beans was studied using liquid chromatography/mass spectrometry. Apart from mycotoxins, the content of ergosterol as a marker indicating fungal occurrence was also determined. Among aflatoxins, aflatoxin B1 was the dominant mycotoxin in coffee bean samples, with the highest level at 17.45 ng g-1 . Ochratoxin A was detected in four samples at levels ranging from 1.27 to 4.34 ng g-1 , and fungi potentially producing this toxin, namely Aspergillus oryzae, Alternaria sp., Aspergillus foetidus, Aspergillus tamarii and Penicillium citrinum, were isolated. CONCLUSION: Steaming and decaffeination of coffee beans increased antioxidant activities of brews in comparison with those prepared from unprocessed beans. Although toxins can be quantified in green coffee beans and novel fungi were isolated, their concentrations are acceptable according to legal limits. © 2017 Society of Chemical Industry.

Experimental Studies on Some Immunotoxicological Aspects of Aflatoxins Containing Diet and Protective Effect of Bee Pollen Dietary Supplement.

Aflatoxins (AFs), widely distributed food-borne mycotoxins, affect quality and safety of food and cause economic losses in livestock. In this study, the protective effect of Bee Pollen (BP) against some immunotoxic hazards elucidated from eating of AFs-containing diet was investigated in Wistar rats. Rats were randomly classified intofour groups and treated for 30 days, Group 1; control negative, Group 2; Total AFs (3 mg kg(-1) basal diet), Group 3; BP (20 g kg(-1) basal diet) and Group 4; AFs+BP in basal diet. The immunoprotective effect of BP was revealed in terms of increasing (relative to levels seen in Group 2 rats that consumed the AFs diet) serum total protein and globulin levels, restored normal neutrophil (PMN)/lymphocyte ratio, increased PMN phagocytic activity and increased lymphocyte proliferative capacity. Also, the use of the BP reduced spleen H2O2 levels and increased GSH content while maintaining normal levels of NO formation. Histopathologic analysis showed thatthe AFs caused lymphocytic depletion in the spleen; however, BP induced lymphocytic hyperplasia and reduced the levels of AFs-inducible cellular exhaustion or depletion. These results provide evidence of a protective effect of BP against some immunotoxic actions induced in situ by consumption of AFs.

Efficacy of Mentha spicata essential oil in suppression of Aspergillus flavus and aflatoxin contamination in chickpea with particular emphasis to mode of antifungal action.

The present study reports in vivo antifungal and antiaflatoxigenic efficacy of Mentha spicata essential oil (EO) against toxigenic Aspergillus flavus strain LHP(C)-D6 in chickpea food system up to 12 months of storage. In addition, the mode of antifungal action of EO was also determined to understand the mechanism of fungal growth inhibition. The in vivo study with different concentrations of M. spicata EO showed dose-dependent decrease in fungal colony count as well as aflatoxin B1 concentration. The EO caused >50% protection in inoculated sets and >70% protection in uninoculated sets of chickpea food system against A. flavus at 1.0 µL mL(-1) air concentration. However, at the same concentration, EO caused 100% inhibition to aflatoxin B1 production in both sets when analyzed through high-performance liquid chromatography (HPLC). The antifungal target of EO in fumigated cells of A. flavus was found to be the plasma membrane when analyzed through electron microscopic observations and ions leakage test. The EO fumigated chickpea seeds showed 100% seed germination and seedling growth after 12 months of storage. Based on these observations, M. spicata EO can be recommended as plant-based preservative for safe protection of food commodities during storage conditions against fungal and most importantly mycotoxin contaminations.

Effect of supplementation of fermented milk drink containing probiotic Lactobacillus casei Shirota on the concentrations of aflatoxin biomarkers among employees of Universiti Putra Malaysia: a randomised, double-blind, cross-over, placebo-controlled study.

Human exposure to aflatoxin is through the diet, and probiotics are able to bind aflatoxin and prevent its absorption in the small intestine. This study aimed to determine the effectiveness of a fermented milk drink containing Lactobacillus casei Shirota (LcS) (probiotic drink) to prevent aflatoxin absorption and reduce serum aflatoxin B1-lysine adduct (AFB1-lys) and urinary aflatoxin M1 concentrations. The present study was a randomised, double-blind, cross-over, placebo-controlled study with two 4-week intervention phases. In all, seventy-one subjects recruited from the screening stage were divided into two groups--the Yellow group and the Blue group. In the 1st phase, one group received probiotic drinks twice a day and the other group received placebo drinks. Blood and urine samples were collected at baseline, 2nd and 4th week of the intervention. After a 2-week wash-out period, the treatments were switched between the groups, and blood and urine samples were collected at the 6th, 8th and 10th week (2nd phase) of the intervention. No significant differences in aflatoxin biomarker concentrations were observed during the intervention. A within-group analysis was further carried out. Aflatoxin biomarker concentrations were not significantly different in the Yellow group. Nevertheless, ANOVA for repeated measurements indicated that AFB1-lys concentrations were significantly different (P=0·035) with the probiotic intervention in the Blue group. The 2nd week AFB1-lys concentrations (5·14 (SD 2·15) pg/mg albumin (ALB)) were significantly reduced (P=0·048) compared with the baseline (6·24 (SD 3·42) pg/mg ALB). Besides, the 4th week AFB1-lys concentrations were significantly lower (P<0·05) with probiotic supplementation than with the placebo. Based on these findings, a longer intervention study is warranted to investigate the effects of continuous LcS consumption to prevent dietary aflatoxin exposure.

Controlling Aspergillus flavus and Aspergillus parasiticus growth and aflatoxin production in poultry feed using carvacrol and trans-cinnamaldehyde.

Aflatoxins (AF) are toxic metabolites primarily produced by molds, Aspergillus flavus and Aspergillus parasiticus. Contamination of poultry feed with AF is a major concern to the poultry industry due to severe economic losses stemming from poor performance, reduced egg production, and diminished egg hatchability. This study investigated the inhibitory effect of 2 generally regarded as safe (GRAS), natural plant compounds, namely carvacrol (CR) and trans-cinnamaldehyde (TC), on A. flavus and A. parasiticus growth and AF production in potato dextrose broth (PDB) and in poultry feed. In broth culture, PDB supplemented with CR (0%, 0.02%, 0.04% and 0.08%) or TC (0%, 0.005%, 0.01% and 0.02%) was inoculated with A. flavus or A. parasiticus (6 log CFU/mL), and mold counts and AF production were determined on days 0, 1, 3, and 5. Similarly, 200 g portions of poultry feed supplemented with CR or TC (0%, 0.4%, 0.8%, and 1.0%) were inoculated with each mold, and their counts and AF concentrations in the feed were determined at 0, 1, 2, 3, 4, 8, and 12 weeks of storage. Moreover, the effect of CR and TC on the expression of AF synthesis genes in A. flavus and A. parasiticus (aflC, nor1, norA, and ver1) was determined using real-time quantitative PCR (RT-qPCR). All experiments had duplicate samples and were replicated 3 times. Results indicated that CR and TC reduced A. flavus and A. parasiticus growth and AF production in broth culture and chicken feed (P<0.05). All tested concentrations of CR and TC decreased AF production in broth culture and chicken feed by at least 60% when compared to controls (P<0.05). In addition, CR and TC down-regulated the expression of major genes associated with AF synthesis in the molds (P<0.05). Results suggest the potential use of CR and TC as feed additives to control AF contamination in poultry feed.

Influence of the antimicrobial compound allyl isothiocyanate against the Aspergillus parasiticus growth and its aflatoxins production in pizza crust.

Aflatoxins (AFs) are secondary metabolites produced by different species of Aspergillus, such as Aspergillus flavus and Aspergillus parasiticus, which possess mutagenic, teratogenic and carcinogenic activities in humans. In this study, active packaging devices containing allyl isothiocyanate (AITC) or oriental mustard flour (OMF) + water were tested to inhibit the growth of A. parasiticus and AFs production in fresh pizza crust after 30 d. The antimicrobial and anti-aflatoxin activities were compared to a control group (no antimicrobial treatment) and to a group added with commercial preservatives (sorbic acid + sodium propionate). A. parasiticus growth was only inhibited after 30 d by AITC in filter paper at 5 µL/L and 10 µL/L, AITC sachet at 5 µL/L and 10 µL/L and OMF sachet at 850 mg + 850 µL of water. However, AFs production was inhibited by all antimicrobial treatments in a dose-dependent manner. More importantly, AITC in a filter paper at 10 µL/L, AITC sachet at 10 µL/L, OMF sachet at 850 mg + 850 µL of water and sorbic acid + sodium propionate at 0.5-2.0 g/Kg completely inhibited AFs formation. The use of AITC in active packaging devices could be a natural alternative to avoid the growth of mycotoxinogenic fungi in refrigerated bakery products in substitution of common commercial preservatives.