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Beneficial and Adverse Effects of Natural, Bioactive Dietary Chemicals on Human Health and Food Safety (From W1122)

Investigators
Romagnolo, Donato
Institutions
University of Arizona
Start date
2007
End date
2007
Objective
  1. Consumption of food-borne bioactive compounds can protect against human diseases such as cancer, inflammation, birth defects, and microbial infection. We will determine the mechanisms by which selected compounds exert their protective action.
  2. Food-borne toxins and carcinogens are present per se or are induced by processing, preparation, and other post-harvest steps. We will identify mechanisms of action and develop biomarkers of natural and induced toxicants in food for human risk assessment and disease prevention.
  3. Selected classes of bioactive compounds show potential for beneficial or adverse effects on human health. We will discover bioactive compounds that have beneficial or adverse effects on human health.
More information
NON-TECHNICAL SUMMARY: Besides genetic changes, epigenetic alterations such as histone and DNA modifications have been proposed to play a key role in the onset of a variety of tumors by inducing stable, heritable changes in gene expression. These changes can occur randomly or in response to the environment. For example, in sporadic breast tumors the BRCA-1 promoter is hypermethylated and the expression of BRCA-1 is reduced in the absence of mutations in the BRCA-1 gene. This suggests that hypermethylation caused by environmental signals could participate in establishing and maintaining heritable modifications of BRCA-1 expression that do not involve mutations. This project examines whether epigenetic events reduce the expression of the tumor suppressor gene, BRCA-1 in breast epithelial cells.

APPROACH: Diet is the most important vehicle of exposure to environmental ligands of the aromatic hydrocarbon receptor (AhR), including dioxins and polycyclic aromatic hydrocarbons (PAHs). The possible role of AhR ligands in the etiology of breast cancer is supported by the following evidence: 1. PAHs such as dimethylbenz[a]anthracene (DMBA) preferentially induce mammary tumors in experimental animals; 2. The disproportionate presence of PAH:DNA adducts in breast tissues from breast cancer patients; and 3. The ability of PAHs to repress BRCA-1 expression in human mammary tumor cell lines. Moreover, prenatal exposure to the dioxin-like compound TCDD (2,3,7,8,-tetrachlorodibenzo-p-dioxin) increased the susceptibility to the development of mammary cancers.

Aim 1 will investigate the effects of AhR ligands on chromatin organization and DNA methylation of the BRCA-1 gene and protective effects of the dietary AhR antagonist resveratrol (RES). Aim 2 will examine in a cell culture model the DMBA-induced histone modifications and DNA methylation at the BRCA-1 promoter and determine if these effects can be reversed by RES. Study Design. For Aim 1, we will use in cell culture conditions breast cancer (MCF-7) cells and normal human mammary epithelial cells. This approach will provide the opportunity to compare the responsiveness of normal vs breast cancer cells to epigenetic regulation of BRCA-1 expression by AhR ligands and the preventative effects of RES. For Aim 2, we will use celll culture as per Aim 1. The treatment with DMBA will occur once two weeks after the initial daily administration of RES. Data acquired through the execution of the proposed experiments will provide 1) a new understanding of the chromatin remodeling events induced by AhR ligands at the BRCA-1 gene both in vitro and in vivo, and 2) provide the foundation for developing preventive strategies against AhR agonists commonly present in the human diet. Because loss of BRCA-1 predisposes to breast cancer development, this work will have a significant impact on understanding how food constituents and natural nutrients help to promote or prevent breast cancer development.

PROGRESS: 2007/01 TO 2007/12
OUTPUTS: The work reported is an outcome of the undergraduate research project of Kristen Scheckel, undergraduate student enrolled in The Undergraduate Research Program of The Honors College at The University of Arizona. This work was presented at the @1st Annual Undergraduate Research Forum. Data are also scheduled to be presented in a poster format at the 2008 Experimental Biology/American Association for Nurition Meetings in San Diego, CA.
PARTICIPANTS: The individuals who participated in the project were Kristen Scheckel, Undergraduate student, who received an Undergraduate Research Training Award as part of the Undergraduate Research Program; Stephanie Degner, who received her Ph.D. in Nutritional Sciences; Corrina Fuentes, undergraduate student and lab assistant; Donato F Romagnolo, Associate Professor and Principal Investigator.
TARGET AUDIENCES: Target audiences for this work include medical and nutrition professionals, graduate students and medical practitioners, and the general public. Specific targets include individuals who may have increased susceptibility to colon and breast cancer, and are interested in learning more about how dietary agents can help in preventing inflammatory processes that increase the risk of colon and breast cancer.
PROJECT MODIFICATIONS: No project modifications are necessary.

IMPACT: 2007/01 TO 2007/12
Mechanisms of prevention of COX-2 activation by the dietary agent rosmarinic acid. One mechanism through which bioactive food components may exert anti-cancer effects is by reducing the expression of the proinflammatory gene cyclooxygenase-2 (COX-2), which has been regarded as a risk factor in tumor development. Rosmarinic acid (RA) is a phenolic derivative of caffeic acid present in rosemary (Rosmarinus officinalis). Previous research documented that RA may exert anti-inflammatory effects. However, the mechanisms of action of RA on COX-2 expression have not been investigated. We report that in colon cancer HT-29 cells, RA reduced the 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced COX-2 promoter activity and protein levels. In HT-29 and breast cancer MCF-7 cells, the cotreatment with RA reduced TPA-induced transcription from a control AP-1 promoter-luciferase construct and binding of the AP-1 factors c-Jun and c-Fos to COX-2 promoter oligonucleotides harboring a cAMP-response element (CRE). The anti-AP1 effects of RA were also examined in a non-malignant breast epithelial cell line (MCF10A) in which RA antagonized the stimulatory effects of TPA on COX-2 protein expression, the recruitment of c-Jun and c-Fos to the COX-2/CRE oligonucleotides, and activation of the extracellular signal-regulated protein kinase-1/2 (ERK1/2), a member of the mitogen-activated protein kinase (MAPK) pathway. Thus, we propose that RA may be an effective preventative agent against COX-2 activation by AP-1 inducing agents in both cancer and non-malignant cells.

PROGRESS: 2006/01/01 TO 2006/12/31
Environmental contaminants found in industrial pollution, tobacco smoke, and cooked foods include benzo[a]pyrene (B[a]P) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), which have been shown to act as endocrine disruptors and tumor promoters. In previous studies, we documented that estrogen (E2)-induced BRCA-1 transcription required the recruitment of an AP-1/ER complex to the proximal BRCA-1 promoter. Here, we report that activation of BRCA-1 transcription by E2 requires occupancy of the BRCA-1 promoter by the unliganded aromatic hydrocarbon receptor (AhR). The stimulatory effects of E2 on BRCA-1 transcription are counteracted by 1) cotreatment with the AhR antagonist 3'methoxy-4'-nitroflavone (3M4NF); 2) transient expression in ER-negative Hela cells of ER lacking the protein-binding domain for the AhR; and 3) mutation of two consensus xenobiotic responsive elements (XRE=5'-GCGTG-3') located upstream of the ER binding region. These results suggest that the physical interaction between the unliganded AhR and the liganded ER plays a positive role in E2-dependent activation of BRCA-1 transcription. Conversely, we show that the AhR ligands B[a]P and TCDD aborgate E2-induced BRCA-1 promoter activity. The repressive effects of TCDD are paralleled by increased recruitment of the liganded AhR and HDAC1, and reduced occupancy by p300 and SCR-1 at the BRCA-1 promoter region flanking the XREs. We propose that the ligand status of the AhR modulates transcriptional activation of the BRCA-1 promoter by estrogen. In a separate study, we investigated the effects of CLA on COX-2 transcription in breast cancer cells. Results of transient transfection studies revealed that treatment of breast cancer MCF-7 cells with a CLA mix or selected isomers (c9, t11-CLA; t10, c12-CLA) at concentrations ranging from 20 to 80 Æ’Ýmol/L, attenuated in a dose-dependent fashion COX-2 transcription induced by the proinflammatory agent 12-O-tetradecanoylphorbol-13-acetate (TPA). In addition, the CLA mix inhibited TPA-induced activity of the human collagenase-1 promoter. Because TPA activates the activator protein-1 (AP-1) transcription factor, we investigated the effects of CLA on recruitment of nuclear proteins to a cAMP response element (CRE) (-59/-53) in the COX-2 promoter and a consensus TPA-responsive element (TRE) in the human collagenase-1 promoter, which are binding sites for AP-1 members. Using electrophoretic mobility shift assays, we found that the CLA mix reduced TPA-induced recruitment of nuclear proteins to the COX-2 CRE and human collagenase-1 TRE. Binding studies revealed that the t10, c12-CLA isomer was more effective than the CLA mix or c9, t11-CLA in reducing binding of cJun to either the COX-2 CRE or human collagenase-1 TRE, whereas LA increased binding to both elements. Overexpression of the AP-1 member, c-Jun, reversed the inhibitory effects of the CLA mix on COX-2 transcription, and restored binding of nuclear proteins to the CRE and TRE. We conclude that the ability of CLA to inhibit AP-1 may contribute to its effectiveness as an anti-carcinogenic and anti-inflammatory agent.

IMPACT: 2006/01/01 TO 2006/12/31
This work will impact our understanding of the dietary risk factor in the etiology of sporadic breast cancer and inflammation and the potential role of preventative dietary agents

PROGRESS: 2005/01/01 TO 2005/12/31
Diet is a vehicle of genotoxic agents, which have been shown to be carcinogenic in experimental models. Our efforst have focused on tumorigenicity induced by xenobiotics such as PAHs that may interfere with expression of tumor suppressor genes such BRCA-1 and p53. Under normal conditions, these tumor suppressors play a key role in control of cell cycle progression and DNA repair. Dysregulation of these genes may increase genomic instability predisposing to tumor growth. We also examined the overlapping effects of PAHs and dioxins on expression of P450s and COX-2 and the role of dietary fatty acids as risk modifiers by influencing the production of proinflammatory and mutagenic endproducts. Finally, we investigated the potential benefits of polyphenolic and synthetic compounds as antagonists of the activated aromatic hydrocarbon receptor (AhR). Whereas the exposure to dietary xenobiotics may be practically unavoidable, future investigations should examine the effects of synergistic interactions between nutrients and dietary xenobiotics on expression and function of tumor susceptibility genes and the physiological role of protective nutritional factors. These efforts may lead to the development of combinatorial dietary interventions to retard and possibly prevent cancer progression.

IMPACT: 2005/01/01 TO 2005/12/31
Interactions between dietary xenobiotics and nutrients influence cancer risk. Epidemiological studies in humans suggested that diet is an important vehicle of exposure to various xenobiotics, including polycyclic aromatic hydrocarbons (PAHs), dioxins, and chlorinated hydrocarbons (CHs). The activation of the aromatic hydrocarbon receptor (AhR) pathway by PAHs and dioxins stimulates the expression of several genes including cytochrome P450s, which metabolize PAHs to highly mutagenic compounds that cause fixation of mutations in the p53 gene and repress the expression of the tumor suppressor gene, BRCA-1. Conversely, PAHs and dioxins have been shown to activate the expression of cyclooxygenase-2 (COX-2), whose protein product participates in the production of reactive PAH-metabolites and synthesis of proinflammatory prostaglandins (PGs). The carcinogenicity of CHs has been attributed to their ability to activate the expression of oncogenes including c-myc, and fatty acid activation of the peroxisome proliferator-activated receptor-gamma (PPARgamma). Dietary components that may protect against the activation of the AhR pathway include flavonoids, which comprise a large family of dietary phenolic phytochemicals found in fruits and vegetables. Research indings produced in our laboratory document the protective effects of natural (genestein) and synthetic (alpha-naphthoflavone) flavonoids against PAH-induced changes in gene expression.

PROGRESS: 2004/01/01 TO 2004/12/31
The environmental carcinogen benzo[a]pyrene (B[a]P) is a prototype polycyclic aromatic hydrocarbon (PAH) found in foods and tobacco smoke, and ligand of the aromatic hydrocarbon receptor (AhR). Conversely, flavonoids compete with PAHs for binding to the AhR. In this study, we wished to characterized in breast epithelial cells the toxicogenomic responses induced by B[a]P and the protective effects of the synthetic flavonoid and AhR-ligand alpha-naphthoflavone (ANF). Acute exposure of breast cancer MCF-7 cells to B[a]P induced a number of changes in physiological state including S-phase arrest, segregation of nucleolar material, and disruption of ribosomal transcription. These changes were antagonized by cotreatment with ANF. Using a cDNA microarray approach, we examined the effects of B[a]P and ANF on the mRNA profile of selected categories of genes encoding for members of the P450 family, DNA damage and repair, and cell cycle regulation. Transcripts that were upregulated by B[a]P included members of the cytochrome P450 family (CYP1A1, CYP1B1, CYP2J2), and cell cycle regulators (cdc28 protein kinase-2). In contrast, B[a]P downregulated transcript levels of glutathione G-transferases (GSTT1, GSTA2, GSTM2), cyclins (A2, D1, and E1), cdc25C, and DNA repair genes including PARP and RCC5. Treatment with ANF alone increased GSTM2 mRNA whereas ANF plus B[a]P tended to restore normal expression profiles. The S-phase arrest induced by B[a]P was accompanied by accumulation of p53 and p21 protein. Conversely, the co-treatment with ANF restored normal cell cycle distribution and prevented the accumulation of p53 and p21. These results highlight important cellular and molecular effects of B[a]P which may be useful biomarkers of exposure to PAHs and the potential usefulness of flavonoids as antagonists of dietary and environmental AhR ligands.

IMPACT: 2004/01/01 TO 2004/12/31
Identify environmental and dietary risk factors in the onset on carcinogenesi of the breast and colon

PROGRESS: 2003/01/01 TO 2003/12/31
Role of Bile Acids and PAHs in Carcinogenesis of the Colon. This project deals with the role of bile acids in the onset of colon carcinogenesis, interactions between bile acids and dietary carcinogens and molecular changes that may predispose to the onset of colon cancer. Deoxycholate, a bile salt present at high levels in the colonic lumen of individuals on a high fat diet, is a promoter of colon cancer. Deoxycholate also causes DNA damage. BRCA1 functions in repair of DNA and in induction of apoptosis. We show that when cultured cells of colonic origin are exposed to deoxycholate at different concentrations, BRCA1 expression is induced at a low non-cytotoxic concentration (10 uM), but is strongly inhibited at higher cytotoxic concentrations (100 uM). Indication of phosphorylation of BRCA1 by DOC (100 uM) at a lower dose was seen by Western Blot analysis, while at a higher dose, DOC (200 and 300 uM) caused a complete loss of BRCA1 expression. We show that BRCA1 is substantially lower in colon adenocarcinomas from five patients compared to associated non-neoplastic colon tissue from the same patients, suggesting that the loss of BRCA1 expression contributes to the malignant phenotype. In the non-neoplastic colon tissue, BRCA1 was localized to the non-goblet cells. Transcriptional and Post-transcriptional Changes Induced by the Carcinogen Benzo[a]pyrene: Effects of the Synthetic Flavonoid alpha-naphthoflavone. The objective of this study was to characterize in breast epithelial cells the toxicogenomic responses induced by benzo[a]pyrene, a prototype polycyclic aromatic hydrocarbon (PAH) and ligand of the aromatic hydrocarbon receptor (AhR), and the B[a]P-metabolite 7r,8t-dihydroxy-9t,10t-epoxy-7,8,9,10-tetrahydro-benzo[a]pyrene (BPDE). Acute exposure of breast cancer MCF-7 cells to B[a]P induced S-phase arrest, loss of membrane integrity, and ultrastructural changes in cellular architecture including segregation of nucleolar material and disruption of nucleolar fibrillar centers. As a means to assess global changes in gene expression induced by PAHs, we analyzed by cDNA microarray the temporal effects of B[a]P, BPDE, and the AhR agonist/antagonist alpha-naphthoflavone (ANF) on the mRNA levels of 1134 genes (GeneMAPTM CancerArray) subdivided in 16-functionally diverse categories. Transcripts that were upregulated by B[a]P included members of the cytochrome P450 family (CYP1A1, CYP1B1, CYP2J2), cell cycle regulators (cdc28 protein kinase-2), histone 3A (H3F3A), chromatin structure (SWI/SNF), peroxisome proliferator activated receptor-gamma (PPARG)and retinoic acid receptor-alpha (RARA). In contrast, B[a]P downregulated transcript levels for various cyclins (A2, D1, and E1), IGFBP-3, Bcl-2, AP-1 (JunD and Fos), fibroblast growth factor receptor1 (FGFR1), and the estrogen receptor-alpha (ESR1). The treatment with BPDE triggered genotoxic responses similar to those elicited by B[a]P, but at lower doses and earlier time points, whereas the co-treatment with ANF tended to counteract the effects of B[a]P on cell cycle arrest, cellular ultrastructure, and mRNA profiles.

IMPACT: 2003/01/01 TO 2003/12/31
Identify environmental and dietary risk factors in the onset on carcinogenesi of the breast and colon

PROGRESS: 2002/01/01 TO 2002/12/31
The objective of this study was to characterize in breast epithelial cells the toxicogenomic responses induced by benzo[a]pyrene, a prototype polycyclic aromatic hydrocarbon (PAH) and ligand of the aromatic hydrocarbon receptor (AhR), and the B[a]P-metabolite 7r,8t-dihydroxy-9t,10t-epoxy-7,8,9,10-tetrahydro-benzo[a]pyrene (BPDE). Acute exposure of breast cancer MCF-7 cells to B[a]P induced S-phase arrest, loss of membrane integrity, and ultrastructural changes in cellular architecture including segregation of nucleolar material and disruption of nucleolar fibrillar centers. As a means to assess global changes in gene expression induced by PAHs, we analyzed by cDNA microarray the temporal effects of B[a]P, BPDE, and the AhR agonist/antagonist alpha-naphthoflavone (ANF) on the mRNA levels of 1134 genes (GeneMAPTM CancerArray) subdivided in 16-functionally diverse categories. Transcripts that were upregulated by B[a]P included members of the cytochrome P450 family (CYP1A1, CYP1B1, CYP2J2), cell cycle regulators (cdc28 protein kinase-2), histone 3A (H3F3A), chromatin structure (SWI/SNF), peroxisome proliferator activated receptor-gamma and retinoic acid receptor-alpha. In contrast, B[a]P downregulated transcript levels for various cyclins (A2, D1, and E1), IGFBP-3, Bcl-2, AP-1 (JunD and Fos), fibroblast growth factor receptor-1 (FGFR1), and the estrogen receptor-alpha. The treatment with BPDE triggered genotoxic responses similar to those elicited by B[a]P, but at lower doses and earlier time points, whereas the co-treatment with ANF tended to counteract the effects of B[a]P on cell cycle arrest, cellular ultrastructure, and mRNA profiles. Changes in the cDNA microarray profiles of a subset of genes following exposure to B[a]P and BPDE were validated by RT-PCR analysis.

IMPACT: 2002/01/01 TO 2002/12/31
Overall, these cumulative data were consistent with previous reports documenting the antiproliferative and endocrine-disrupting effects of AhR-ligands. The current studies suggest that acute exposure to PAHs may induce specific adaptive responses in cellular architecture and expression profile, which may be useful in developing predictive biological models of exposure to PAHs.

PROGRESS: 2001/01/01 TO 2001/12/31
The decontamination of aflatoxin B1 (AFB1)-contaminated corn using an integrated ammoniation fermentation process was studied. Different concentrations [0.25, 0.50, 1.00, 1.5, and 2.0% (w/w)] of ammonium persulfate were tested in the detoxification of AFB1 contaminated corn during fermentation. In order to increase the ethanol production during fermentation and decontamination of corn, 0.5 and 1.0 % (w/w) of azodicarbonamide, benzoyl peroxide, or hydrogen peroxide were tested. Peroxides were added at three different stages of the fermentation process: liquefaction, saccharification, and fermentation. Levels of AFB1 and ethanol were determined after every fermentation process. Treated corn was tested for mutagenic potential using the Salmonella/microsomal mutagenicity assay with TA 100 tester strain and pure AFB1 as positive control. The greater level of decontamination was obtained after the addition of 2.0% (w/w) ammonium persulfate without any effect on the ethanol production. The addition of peroxides did not significantly increase the ethanol production or significantly improve the decontamination process. The best process for decontamination of corn included the addition of 2.0% (w/w) ammonium persulfate and 1.0% (w/w) benzoyl peroxide. The best process for ethanol production was the addition of 2.0% (w/w) ammonium persulfate and 0.5% (w/w) benzoyl peroxide. No treated corn samples showed mutagenic potential.

IMPACT: 2001/01/01 TO 2001/12/31
This process could be used for the production of fuel ethanol from aflatoxin containing corn and rendering the by-product, distillers dried grains and solubles, free from aflatoxin.

PROGRESS: 2000/01/01 TO 2000/12/31
Treatment of whole and ground corn naturally contaminated with 200 ppb total aflatoxin with stabilized granulated ammonium bicarbonate at 25 or 37 degrees, at 10, 15, or 17.5% moisture levels for 0 to 20 days with samples taken at 5 day intervals was unsuccessful in lowering levels of the mycotoxin under any of the conditions described. We are continuing the study using regular powdered ammonium bicarbonate and subjecting the samples to the same testing parameters. We expect that release of ammonia will be more rapid and destruction of the aflatoxin will be similar to our previous extrusion experiments.

IMPACT: 2000/01/01 TO 2000/12/31
Continued investigation into methods of elimination of aflatoxin and fumonisin from already-contaminated commodities may provide an economical means to avoid the destruction of that product and at the same time assure safety to those consuming it.

PROGRESS: 1999/01/01 TO 1999/12/31
We are following the production of fumonisin in culture to see if it is first produced and then destroyed by substances within the culture itself. If so, we want to isolate the substances responsible. Four flasks containing sterile nutrient broth were inoculated with a suspension of a fumonisin- producing strain of Fusarium moniliforme. All were held at 25 degrees C, two flasks with agitation and two without. Samples taken from the flasks for each of 27 days were analyzed for fumonisin. Fumonisin was detectable in the agitated flasks after 3 days with maximum levels at 7 days. Then fumonisin was gradually reduced until day 14 after which a drastic reduction took place. This would indicate that formed fumonisin was being destroyed by some factor in the media, perhaps by the aged mold culture. Cultures in the non-agitated flask and in flasks with moistened, inoculated corn meal and whole corn did not produce fumonisin under our experimental conditions. Whole and ground corn were treated with granulated ammonium bicarbonate at 25 or 37 degrees, at 10, 15, or 17.5% moisture levels for 0 to 20 days with samples taken at 5 day intervals. This study was to find a simple, inexpensive way to treat high moisture corn on the farm while awaiting the drying process. Samples were extracted with either water/methanol (20/80)or 0.1 N HCl/methanol (20/80) to verify the complete destruction of the aflatoxin. Treatment of the whole grain were largely unsuccessful in reducing aflatoxin. We are analyzing the ground corn. It is possible that our use of a stabilized ammonium bicarbonate, normally used for soil treatment, did not allow the development of sufficient ammonia to penetrate the kernels.

IMPACT: 1999/01/01 TO 1999/12/31
Continued investigation into methods of elimination of aflatoxin and fumonisin from already-contaminated commodities may provide an economical means to avoid the destruction of that product and at the same time assure safety to those consuming it.

PROGRESS: 1998/01/01 TO 1998/12/31
We are attempting to monitor the production of fumonisin in synthetic culture and in corn to see if it is first produced and then destroyed by substances within the culture itself as is aflatoxin. If this is so, we want to isolate the substances responsible. Four flasks containing sterile nutrient broth were inoculated with a suspension of a fumonisin-producing strain of Fusarium moniliforme. All were held at 25 degrees C, two flasks with agitation and two without. Samples taken from the flasks for each of 27 days were analyzed for fumonisin. Fumonisin was detectable in samples taken from the agitated flasks after 3 days with maximum levels at 7 days. After that levels were gradually reduced until day 14 after which a drastic reduction took place. Fumonisin levels were close to zero for the rest of the experiment. This would indicate that formed fumonisin was being destroyed in the latter part of the experiment by some factor in the media, perhaps by the aged mold culture. Cultures in the non-agitated flask and in flasks with moistened, inoculated corn meal and whole corn did not produce fumonisin under our experimental conditions

PROGRESS: 1997/01/01 TO 1997/12/31
The project under this objective is in its beginning stages. We are attempting to monitor the production of fumonisin in synthetic culture to see if it is first produced and then destroyed by substances within the culture itself as is aflatoxin. If this is so, we want to isolate the substances responisible. We have been collecting preliminary information, learning HPLC techniques for fumonisin analysis and trying to obtain a fumonisin producing strain of Fusarium. A survey of commercially produced corn-based snack products has shown no fumonisin present.

PROGRESS: 1996/01 TO 1996/12
Fumonisin (FB1)-contaminated corn during nixtamalization w/ Ca(OH)2 treated w/ NaHCO3 + H2O2, w/ NaHCO3 or w/ H2O2 alone, gave 100% reduction of FB1. Brine shrimp mortality w/ 0.25 g FB1 /reaction products dropped from 40% (untreated corn extract) to 17% (chemically-treated corn). FB1 concentrations of 0.0025-100 g/plate showed a small pos. mutagenicity. No newly formed mutagens were detected in treated samples. Untreated corn extract exhibited positive mutagenicity; subsequent analysis showed 65 ppb of aflatoxin B1 which would account for the above mutagenicity. Aflatoxin-contaminated corn containing 140 g/kg was fermented with 2% ammonium persulfate and H2O2, azocarbonamide, or benzoyl peroxide added at 2 concentrations (0.5 or 1%)at different stages (liquefaction, saccharification & fermentation) of the process.The most significant aflatoxin decrease was in samples treated with 2% ammonium persulfate and 0.5% benzoyl peroxide (97%). The highest ethanol production was found in samples treated with 2% ammonium persulfate with 0.5% benzoyl peroxide added during saccharification just prior to fermentation inoculation.The same treatment with 1% benzoyl peroxide inhibited ethanol production.All samples tested negative using the Salmonella/mammalian microsomal mutagenicity assay with TA100. Samples of the original corn combined with pure AFB1 standard showed a decrease in mutagenic activity. This suggests the presence of compound(s) in corn which interfere with the mutagenic activity of aflatoxin.

PROGRESS: 1995/01 TO 1995/12
Nixtamalization of fumonisin B1 (FB1 )-contaminated corn using Ca(OH)2 and NaHCO3 + H2O2 or H2O2 alone gave 100% reduction of FB1. Brine shrimp mortality with FB1/reaction products dropped from 40% to 17%. No newly formed mutagens were detected in the treated samples. Fumonisin B1 in acetic acid/sodium acetate buffer was destroyed by NaOH, NH4OH, and (NH4)2S2O8 and decreased by hydrogen (96%) and horseradish peroxide (70%). Toxicity of samples to brine shrimp was equal to or greater than that of the fumonisin control. Selected Arizona cotton fields were monitored for A. flavus contamination at different heights of the cotton plant and for meteorological factors using the AZMET program. Aflatoxin was determined in immature cotton bolls and cottonseed prior to harvest and in modules. There were higher levels of A. flavus on lower portions of the plant. Aflatoxin contamination was 5 times higher on insect damaged cotton bolls than on normal bolls. High relative humidity and rainfall favored A. flavus contamination. Aflatoxin-contaminated roasted peanuts cooked in water with hydrogen peroxide in combination with sodium or ammonium bicarbonate, sodium hydroxide, and ammonium persulfate had aflatoxin reduced by half by cooking alone. None of the chemicals alone or in combination with hydrogen peroxide were successful in eliminating aflatoxin from the peanut sauce.

PROGRESS: 1994/01 TO 1994/12
Glutathione (GSH) added to the Salmonella mutagenicity assay decreased the mutagenic potential of aflatoxin B1 (AFB1) AFB2 and AFB1:AFB2 mixtures to TA100. S9 mix converted about 5% of AFB2 to AFB1. Aflatoxins were broken down by lactoperoxidase (LP) and horseradish peroxidase (HRP)with oxygen present, with low H202 concentrations (0.1-1%), but not in the absence of oxygen or H202. In vitro fumonisin B1 was reduced 70-85% by treatment with 0.01M Ca(OH)2, 3% H202 , and 2% Na2CO3 alone or in combinations for 0.5 to 12 hours at 23C. Optimum breakdown was with a combination of all reagents, but including peroxide in the reaction mixture had little addtional effect. Brine shrimp and microbiological assays confirmed the reduction of fumonisin. Extracts from aflatoxin- free corn and aflatoxin- contaminated corn with and without ammonia treatment, tested for mutagenic potential using the Ames test (TA 100) gave evidence of an "anti aflatoxin mutagenic factor" in corn that may block the mutagenic activity of aflatoxin in the Ames test.

PROGRESS: 1993/01 TO 1993/12
Naturally-contaminated, roasted peanuts containing with 2600 ppb total aflatoxin(AF) were ground and cooked with water, H(subscript 2)O(subscript 2), NaHCO(subscript 3), NH(subscript 4)HCO(subscript 3), or ammonium persulfate. Cooking alone reduced AF by half. Although cooking with food additives further reduced AF, no chemical alone or in combination with H(subscript 2)O(subscript 2) were successful in eliminating AF. Naturally-contaminated corn containing 500 ppb total AF was fermented using commercial procedures to determine fate of aflatoxin and alcohol production. H(subscript 2)O(subscript 2), NaOH, NH(subscript 4)OH, NH(subscript 4)Cl, ammonium persulfate, catalase, and horseradish peroxidase alone or in combinations were added during fermentation. Alcohol production was lower with NH(subscript 4)Cl but higher when catalase and horseradish peroxidase were used together. High levels of H(subscript 2)O(subscript 2) produced slightly more alcohol but slowed or inhibited fermentation. Aflatoxin was reduced in DDGS of samples treated with either ammonium or sodium hydroxide. H(subscript 2)O(subscript 2), catalase, and horseradish peroxidase alone caused a reduction in AF; greatest reductions occurred with the combination of H(subscript 2)O(subscript 2), catalase, and peroxidase. Ammonium persulfate destroyed nearly all aflatoxin, but completely inhibited production of alcohol.

PROGRESS: 1992/01 TO 1992/12
Naturally-contaminated corn with 500 ppb and in-shell peanuts with 280 ppb totalaflatoxin (AF) with 10, 15, 17.5 or 20% moisture were stored in plastic bags at 21(degree), 32(degree), or 43(degree) C for 5, 10, 20, or 40 days. There was no increase in AF in corn with moisture of 10, 15 or 17.5%. There was an increase in toxin after 10, 20 or 40 days at 20% moisture with levels dependent upon temperature. AF in corn stored at higher temperatures decreased after 20 days. AF in peanuts stored in the shell increased under all storage conditions. Analysis of tortillas made from the above corn with 0.75% Ca(OH)(subscript 2) and 0, 0.1, 0.5, or 1% ammonium bicarbonate during cooking indicated that those made using ammonium bicarbonate had less AF than those which did not. No process decreased the AF to below the 20 ppb level except for the 1% ammonium bicarbonate sample (13 ppb). The above corn was fermented using NaOH or NH(subscript 4)OH to increase the pH to 9.6 during liquefaction. H(subscript 2)O(subscript 2), lactoperoxidase and/or horseradish peroxidase were added during the process. Fermentation with NH(subscript 4)OH showed significant decreases in AF which were even greater with the peroxide and peroxidases. Fermentation with NaOH was unsuccessful in reducing AF to below 20 ppb.

PROGRESS: 1991/01 TO 1991/12
All experiments used field corn, naturally-contaminated with 400 ppb aflatoxin (AF). Corn meal was extruded from an X-5 Wenger extruder at temperatures above 100(degree)C with 16% added moisture and combinations of H(subscript 2)O(subscript 2) and NH(subscript 4)OH or NH(subscript 4)HCO(subscript 3). Extrusion alone permanently destroyed about 50% of the AF. Only NH(subscript 4)OH or NH(subscript 4)HCO(subscript 3) reduced AF below 20 ppb. The odor of ammonia could not be sensed on the product afterwards. Neither the original or treated samples produced the expected mutagenicity with the Ames test. No destruction of amino acids by extrusion under alkaline conditions was evident. Corn meal was treated with 1 or 2% H(subscript 2)O(subscript 2) and 1% NaOH with and without preheating (75(degree)C) for 30 min. All treatments were equally successful in permanently lowering the AF by 90%. In no case was AF decreased to below 20 ppb. Tortillas were made from whole corn was cooked for 1.25 h in 0.75% Ca (OH)(subscript 2) (w/w) and allowed to soak 15 hr. Also 0.1, 0.5 or 1% ammonium bicarbonate was added. Using increasing ammonium bicarbonate resulted in lowered AF. No process decreased the AF to below 20 ppb. Whole corn in plastic bags in 500 g lots with 10, 15, 17.5, and 20% total moisture was stored at 70, 90, and 110(degree)C for 5, 10, 20, and 40 days. AF increased drastically only in samples stored at 20% moisture; after 20 days it decreased to below original levels.

PROGRESS: 1990/01 TO 1990/12
Milk obtained from cows fed rations containing aflatoxin-contaminated cottonseed, ammonia-treated aflaxtoxin-contaminated cottonseed, and uncontaminated cottonseed were tested for mutagenic potential using the Ames Salmonella assay. Samples included whole milk, nonfat dry milk powder, cream, and reconstituted whole milk. High mutagenic activity was seen in samples from contaminated cottonseed but not in samples from cows fed the ammonia treated, contaminated cottonseed. Field corn, naturally-contaminated with 400 ppm aflatoxin was extruded above 100C with 16% added moisture and combinations of calcium hydroxide, hydrogen peroxide, sodium bicarbonate, ammonium hydroxide, sodium bisulfite, ammonium bicarbonate, azodicarbonamide, and benzoyl peroxide. Extrusion alone destroyed approximately 50% of the aflatoxin. Ammonium hydroxide and ammonium bicarbonate reduced aflatoxin below 20ppb. Changes in aflatoxin B(1) and B(2) mutagenic potentials were determined using the Salmonella assay with and without S9 liver homogenate and glutathione (GSH). Addition of GSH to the assay showed a decrease in the mutagenic potential for AFB(1), AFB(2) and mixtures. GSH without S9 eliminated the mutagenic potentials of AFB(1) and AFB(2). The AFB(1):AFB(2) mixture of 1:0.3 most closely resembled activity of AFB(1) alone.

PROGRESS: 1989/01 TO 1989/12
Aflatoxin contaminated whole cottonseed was treated with ammonia using two commercial methods, a high pressure, high temperature process (14% moisture, 4% ammonia gas, 100C, 40 lbs/in, 30 min, 1200 mu g/Kg) and an atmospheric pressure, ambient temperature process (1 1/2% anhydrous ammonia, 15% water, ambient temperatures, 42 days, 5200 mu g/Kg). Rations containing the treated seed were fed to dairy cows (15 and 25% of the ration, respectively), and milks obtained were dried and incorporated into trout (Salmo gairdneri) diets (25%). In the pressure method, aflatoxin M(1) in the untreated seed/milk trout diets was 8 mu g/Kg. No tumors nor neoplastic lesions were observed in livers of trout fed milk from either the ammonia-treated or untreated source. In the ambient method, 18% of the trout consuming milk from cows with non-ammoniated cottonseed in the ration had liver tumors; those consuming milk from cows with ammonia-treated aflatoxin-contaminated cottonseed in their rations had no tumors nor neoplastic lesions. These data demonstrate that the ammonia process reduces risks posed by aflatoxin contamination.

PROGRESS: 1988/01 TO 1988/12
Mutagenicity of terminal aflatoxin B(subscript 1) and B(subscript 2) precursors were determined using Ames test (TA 98) in rat liver homogenate. O-Methylsterigmatocystin (OMST) and AFB, exhibited some cytotoxicity to the tester strain. Sterigmatocystin and OMST exhibited positive responses. 1 ug OMST is required to show same response exhibited by 0.02 u2 AFB(subscript 1). AFB(subscript 2) and precursor did not show activity. Maximum levels tested were 45 ug/plate for dehydro-OMST, and 100 ug/plate for hydroxy-OMST and AFB(subscript 2). Effects of glutathione with and without AFB(subscript 2) on AFB(subscript 1) mutagenic potential in the Ames test and TA 100 was determined with and with S9. There was a decrease in potential of both AFB(subscript 2) and AFB(subscript 1), when glutathione was added to S9. AFB(subscript 2) had no effect on potential of AFB(subscript 1) in concentration of 0.3:1. With the 1:1 effect as additive. For plates which contained neither glutathione nor S9, there was a decrease in potential and cytotoxicity. AFB(subscript 1) - contaminated cottonseed (1500 ppb) was ammoniated and fed to cows as 25% of ration. Milk (3 ppb aflatoxin M(subscript 1)) was spray-dried for trout studies. Milks were tested for potential using Salmonella/ microsome mutagenicity strains TA 98 and TA 100. Samples from cows exposed to aflatoxin cottonseed and the spiked milks showed high levels of activity.

PROGRESS: 1987/01 TO 1987/12
Jojoba albumins and globulins were treated with jojoba tannins to study the protein-tannin complex formation. Trypsin and chymotrypsin were also treated with jojoba tannins to determine the change of enzymatic activity. The treated albumins and globulins were fractionated by gel filtration before and after treatment. Two albumin fractions did not appear after treatment but a new fraction appeared with a shorter elution time. A precipitate was formed by addition of jojoba tannins to the jojoba globulins and two globulins fractions did not appear after treatment. Trypsin was completely inhibited by 5.80 mu g of tannin/ml. Chymotrypsin was completely inhibited by 36.44 mu g of tannin/ml. Whole cottonseed, naturally-contaminated with AFB(1) was ammoniated commercially and fed as 25% of a dairy ration to 3 cows. The contaminated whole seed was likewise fed. The cream was removed from the milk of these cows, the skim milk was spray-dried and the powder from both treatments was sent to Oregon State for evaluation of carcinogenicity with rainbow trout.

PROGRESS: 1986/01 TO 1986/12
Naturally-contaminated yellow dent corn containing 1,500 mu g total aflatoxin per kg was treated with 2.5 or 3% NaOH at 100C for 2 or 5 min respectively. After washing, the moisture was adjusted by soaking or autoclaving and the product was deep fried for 10 or 15 min to produce a corn snack. There was greater than 99% reduction of the aflatoxin in the final product. Acidifying products prior to analysis caused reformation of as much as 18% of the original aflatoxin; the acidified final product showed no reformation. Mutagenesis detected by Salmonella typhimurium (TA 98 and TA 100) decreased as aflatoxin levels decreased. Mutagenicity of the final product was equal to that of the negative controls. Naturally-contaminated spray dried milk was added to raw or pasteurized whole milk to a final concentration of 1.1 mu g AFM(1)/1. Formalin (37% w/w) was added to these milk solutions to give concentrations of zero, .025, .05 and 0.1% formaldehyde, and samples were stored in the dark at 21C in either plastic or glass containers. Samples were analyzed for AFM(1) at 0, 1, 2, 3 and 4 weeks. A second trial used only raw milk and glass containers with AFM(1) analyses at 0, 1, and 2 weeks. Twenty-five percent less AFM(1) was detected in milk stored in plastic than in glass containers, but whether the milk was raw or pasteurized had no effect. AFM(1) losses increased during storage and with increasing formaldehyde concentration. AFM(1) in milk preserved with 0.

PROGRESS: 1985/01 TO 1985/12
The effects of glutathione (GSH) and the combination of GSH and glutathione-S-transferase (G-S-T) on aflatoxin B(1) (AFB(1)) mutagenesis using Salmonella typhimurium strains TA 98 and TA 100 were tested. Ten concentrations of AFB(1) (0 to 1.0 ug/plate) were added to a liver microsomal homogenate (S9 mix) and incubated for 0 and 30 minutes at 37 C prior to plating. One third of the samples contained GSH and one third contained G-S-T. Of the samples incubated prior to plating one half contained bacteria. Addition of GSH and GSH+G-S-T decreased both mutagenic activity and lethality of AFB(1) to the bacteria. Formation of the AFB(1)-GSH conjugate was verified using reverse phase HPLC. A simplified procedure for treating AFB(1) contaminated, whole cottonseed in the seed handling system at the gin was tested. Water and anhydrous ammonia were metered into the pneumatic seed handling systems at 10 and 1.5% respectively. Samples taken from the gin stream every 10 minutes for 7 hr each day for 3 days and from each pile after the entire run and at 24, 48, 72, and 96 hr intervals indicated no reduction of AFB(1). Yellow corn which was naturally-contaminated with 1570 ppb of total aflatoxin was treated with 2.5 or 3% NaOH for 2 or 5 min. After washing and moisture adjustment the product was deep fried for 10 or 15 min. Both manufacturing processes were effective in reducing aflatoxin in the finished product to nearly non-detectable levels and mutagenic activity to background.

PROGRESS: 1984/01 TO 1984/12
The effectiveness of cottonseed ammoniation for removal of carcinogenic substances from milk from cows consuming aflotoxin-contaminated whole cottonseed as a part of their diet was determined. Milk was obtained from cows which had consumed aflatoxin-contaminated whole cottonseed (5000 mu g/kg) or the same cottonseed which had been treated with 11/2% anhydrous ammonia and 10% water at ambient temperatures and pressure. These two samples along with appropriate controls were included as 20% of the diet of Rainbow trout for 9-12 months. No hepatomas were found in 2 groups of 100 trout consuming regular trout rations, a control diet containing 20% normal non-fat dried milk, or a diet containing 20% milk from cows consuming ammoniated cottonseed. Hepatomas (26%) were found in trout consuming a control diet with 4 mu g/kg aflatoxin B(1), and in trout (18%) consuming a diet containing 20% milk from cows consuming non-treated, contaminated cottonseed. Aflatoxin M(1) has about 1/3 the carcinogenicity of AFB(1) to trout: ammoniation of the cottonseed reduces carcinogneicity of the trout diets to that of the control. Ninety high producing dairy cows were fed aflatoxin-contaminated whole cottonseed as 15% of their diet for 70 days. Aflatoxin levels in their milk indicated that approximately 1.6% of the dietary aflatoxin is excreted in the milk. Six dairy cows were fed aflatoxin, ammoniated aflatoxin or ammoniated aflatoxin which had been acidified.

PROGRESS: 1983/01 TO 1983/12
Five processes for tortilla manufacture all decreased aflatoxin (AF) from naturally contaminated corn up to 88%; however, acidifying all products prior to analysis caused reformation of up to 44% of the original aflatoxin. Mutagenicity of the acetone extracts was also decreased by tortilla manufacturing processes. Treatments of aflatoxin-contaminated peanut meal with NaHSO(3), H(2)O(2), and NaOC1 with or without 1% Ca(OH)(2) were all effective in reducing aflatoxin levels at 0.5 and 0.2% concentration. Alkaline media greatly reinforced the effectiveness of the oxidizing agents. All treatments were effective in decreasing mutagenic response of S. typhimurium TA 98 and TA 100 strains. Complete elimination of AFB(1) or mutagenicity was not obtained under these treatment conditions. Comparison of results obtained from chemical analyses with those obtained with the Ames test gave a correlation coefficient of 0.9. Direct HPLC injection of the ammonia-AFB(1) reaction media indicated the rapid formation of two very polar by-products. These compounds were stable at pH 12 but reverted quickly to AFB(1) during mass spectral analysis.

PROGRESS: 1982/01 TO 1982/12
Naturally contaminated peanut meal was treated with various chemicals at different concentrations (0, 0.5, and 2.0%) and different temperatures (22 and 62 degrees C), and aflatoxin degradation was measured. All treatments, NaHSO(3), H(2)O(2) plus 1% Ca (OH)(2), NaOCl, and NaOCl plus l% Ca (OH)(2), were effective in decreasing the amount of aflatoxin and the mutagenic response of S. typhimurium TA98 and TA100 strains. Adding 1%Ca(OH)(2) to NaCCl decreased mutagenic activity significantly; whereas adding it to H(2)O(2) did not. Of all treatments, NaOCl plus 1%Ca(OH)(2) was the most effective; nevertheless complete elimination of aflatoxin and of mutagenicity of extracts was not achieved. The higher temperature caused a significant decrease of both aflatoxin and mutagenicity. Comparison of analytical results with results of the Ames test gave a correlation coefficient of 0.9. Crystalline aflatoxin B(1) was treated with concentrated aqueous ammonia at 25 degrees C under aerobic and anaerobic conditions. Samples were removed periodically and tested for residual aflatoxin and reaction products. At least two reaction products with a greater polarity than aflatoxin B(1) were formed. During identification attempts using mass spectrometry, these compounds reverted to aflatoxin B(1). Temperatures higher than 60 degrees C during ammoniation were necessary to cause changes in aflatoxin B(1) which were not reversible under acidic conditions.

PROGRESS: 1981/01 TO 1981/12
Experimental shrimp, Penaeus stylirostrus and P. vannamei, were exposed to 2.0 to 160mg aflatoxin B(1) per kg body weight by intramuscular injection or by multiple per os dosing with feed (53 to 300 mg aflatoxin B(1)/kg feed) for up to 24 days. Histopathogenesis of aflatoxicosis in the exposed animals was time and dose-dependent in the hepatopancreas, mandibular organ, and in the hematopoietic organs. Inconsistant lesions were observed in other organs and tissues. Aflatoxin B(1) was added in various concentrations to skim milk inoculated with three strains of Lactobacillus and three strains of Streptococcus in pure and mixed cultures. Skim milk contaminated naturally with 0.6 Mug aflatoxin M(1)/1 was inoculated with the same lactic acid bacteria. Aflatoxin B(1) caused an increase in the chain length of Streptococcus bacteria but caused no significant changes in morphology nor in coagulation time of the milk. Aflatoxin M(1) delayed coagulation of skim milk by over 33 hours.

PROGRESS: 1980/01 TO 1980/12
Aflatoxin-contaminated cottonseed was treated with 1-1/2% ammonia and 10% water and packed into 10 foot diameter by 100 foot long polyethene bags and held for 21 days. The ammoniated cottonseed was fed to lactating dairy cattle in two different trials. Analysis of milk for aflatoxin M(1) showed a maximum level of 1.8 ppb from the group receiving the untreated cottonseed and a maximum of 0.8 from the group receiving the ammoniated meal. Both groups returned to negative 6 days after cottonseed feeding ceased. Dried citrus waste was fed to dairy cows and their milk was analyzed according to the standard methods for AFM(1). Results indicate that a compound from the citrus waste may interfere with the analysis of aflatoxin M(1) in milk by HPLC. This interference can be overcome by several methods. A short-term toxicity study of aflatoxin B(1) on Benaeid shrimp involving three major routes of contact: 1) intramuscular injection, b) ingestion, and c) absorption through the gills indicated that 0.600 mg aflatoxin B(1) per kg feed did not significantly increase the mortality rate even after three weeks. In addition, little cellular damage was observed. Injection of 51 mg aflatoxin B(1) per kg body weight caused severe damage and mortality after 5 days. No damage was observed in the hepatopancreas from doses less than 2 mg aflatoxin B(1) per kg body weight. There was little lethality due to absorption of aflatoxin B(1) at 10 mg/L seawater.

PROGRESS: 1979/01 TO 1979/12
1. Reduction of aflatoxin M(1) (90%) in milk from cows fed whole cottonseed contiminated with 680 ppb aflatoxin B(1) (AFB(1)) was achieved by treatment of the seed with 1 l/2% aqueous ammonia and 10% added H(2)O. Ammoniation was carriet out in a 10X8X100 ft. long polyethylene bag using an "Ag bagger". 2. Distruction of AFB(1) in whole cottonseed atmospheric pressure was determined at 0, 1, 1.5 and 2% aqueous ammonia, 7.5, 15, 17.5 and 20% moisture, at 70, 90 and 110 degrees F, for 5, 10, and 15 days. The most rigorous treatment reduced AFB(1) from 800 ppb to below 10 ppb. 3. Acute and subacute toxicity effects to AFB(1) upon Peneaid shrimp showed an approximate LD(50) of 30-40 Mug/g in 24 hours and 15-20 Mug/g in 96 hours. Histological effects were observed in the feeding (subacute) study. 4. During separation of AFM(1) naturally contaminated milk into fractions, more than 50% remainder in the whey. Upon ultrafiltration of the whey, 75% of the AFM(1) remained in the permeate. Upon separation of the proteins, 86% of the AFM(1) was lost, 14% remained with the globulin fraction and none followed the albumin fraction.

PROGRESS: 1978/01 TO 1978/12
1) Samples of Arizona commercial milk were shown to have levels of aflatoxin M(1) above 1(ppb) during July and August. Cause was determined to be a large lot of extremely contaminated cottonseed which had entered the dairy feed chain. Analysis of milk produced previous and subsequent to these months had insignificant amounts of aflatoxin (less than 15% of dairies with less than or equal to 0.1 ppb). 2) During ultrfilitration of cheese whey, aflatoxin M(1) from naturally contaminated milk was concentrated in the whey protein fraction. An unknown aflatoxin metabolite which interferes with Af M(1) analysis in both TLC and HPLC has been isolated and determined not be Af M(1), M(2), nor B(2a). R(f) values also eliminate other known Af metabolites. Attempts at identification are continuing. 4) Examination of peanuts, peanut products, and milk from commercial sources near Guaymas, Sonora, Mexico showed all peanut candy and 80% of the peanut butter to exceed 20 ppb aflatoxin B(1). Whole and canned peanuts had much lower levels, and no aflatoxin M(1) was found in milk samples.

PROGRESS: 1977/01 TO 1977/12
Thin layer chromatography was used to separate resorcinol compounds, and Rf values were determined for several wheat and triticale grain samples. Different Rf values were found for wheat and triticale. Jojoba meal was fed to weanling mice at two levels for either 4 or 6 weeks. The mice were removed from the feed, raised to sexual maturity, and mated. Mice fed jojoba meal for 6 weeks at both dietary levels had reduced numbers of pups born and weaned. Paired mice fed at a rate of equal body weight had no impairment of reproduction. Aflatoxin M(1) was found in detectable levels (=0.1 ppb) in 10-15% of bulk tank milk samples collected on the bimonthly basis from 154 different dairies affitrated with United Dairymen of Arizona (UDA). Testing of a large number of dairy feed ingredients in Arizona has implicated cottonseed and cottonseed products as the sources of aflatoxin for contamination of milk. Dairy cows from the University of Arizona herd were fed varying levels of aflatoxim and milk was weighed and tested for aflatoxin. No loss in milk production was observed at the levels fed. Tests of 100 cows from each of five Arizona dairies suggest that cows within a herd being fed the same contaminated mix do not secrete aflatoxin in the milk at the same levels. Evidence points to feed selection and not to differences in metabolism of the aflatoxin by the individual cow. Attempts to produce antibodies to patulin by injecting rabbits with patulin conjugated to ficoll or to epoxysepharose were unsuccessful.

PROGRESS: 1976/01 TO 1976/12
Weanling mice were fed four diets for a period of three weeks and then placed on mouse chow. The dietary treatments included controls, 5% jojoba meal, 10% jojoba meal and purified simmondsin being added at a level equal to the 5% jojoba meal. At maturity, the mice were mated and reproduction records kept. The incorporation of either jojoba meal or purified simmondsin in the diet of weanling mice for a period of three weeks caused reduction in number of pups born and weaned. The isolated simmondsin had results similar to the 5% jojoba meal treatment. The simmondsin of jojoba meal affects the reproduction in mice. The naturally occurring toxin, resorcinol, of grain is being tested chemically for structure. The resorcinol has been demonstrated to lower ME value of grain samples. Grain samples with approximately equal resorcinol concentrations do not have similar ME values. These differences may be do to different resorcinol compounds.

PROGRESS: 1975/01 TO 1975/12
The simmonds in toxin of the jojoba bean was isolated and purified. The purified crystals of simmondsin are to be added to whole egg control diets and fed to weanling mice for a period of three weeks. The mice are to be placed on laboratory chow and mated for a reproduction study. Various grain samples have been tested for resorcinol concentrations. In addition, the standard curve usedin the resorcinol method is being investigated. Future plans call for the use of thin-layer chromatography to identify the various types of resorcinol compounds present.

PROGRESS: 1974/01 TO 1974/12
Jojoba meal was fed at levels of 0, 5 and 10% of the diet by replacing equal amounts of soybean meal and resulted in reduced growth in weanling mice at the 5and 10% level. After the initial four weeks the mice were placed on laboratory chow and paired for breeding purposes. The initial mating of mice resulted in 9, 5 and 2 pups for the average number born to the 0, 5 and 10% treated mice. The additional crossing of the different treated males and females resulted in reduced litter size by the 5 and 10% treated female mice. Resorcinol values were determined for 23 varieties of wheat, 5 varieties of triticale, 6 varietiesof sorghum and 5 varieties of barley grains. Correlation studies are under way to determine the relationship between resorcinol levels of the grain and their protein quality plus metabolizable energy values.

PROGRESS: 1973/01 TO 1973/12
A series of three feeding experiments were run with jojoba meal in both weanlingand young adult mice. The jojoba meal, when fed as the sole source of protein in the diet, resulted

Funding Source
Nat'l. Inst. of Food and Agriculture
Project source
View this project
Project number
ARZT-137074-R-23-100
Accession number
64430
Categories
Mycotoxins
Natural Toxins
Commodities
Produce
Nuts, Seeds