ChemInform Abstract: Pigments of Fungi. Part 69. Total Synthesis of (R)-Ochratoxin α (I) and the Formal Total Synthesis of Ochratoxin A (II).

ChemInform ◽  
2010 ◽  
Vol 33 (45) ◽  
pp. no-no
Author(s):  
Christoph D. Donner ◽  
Melvyn Gill
Keyword(s):  
Total Synthesis ◽  
Ochratoxin A ◽  
Ochratoxin Α

Pigments of Fungi. LXIX. Total Synthesis of (R)-Ochratoxin α and the Formal Total Synthesis of Ochratoxin

2002 ◽  
Vol 55 (3) ◽  
pp. 213 ◽  
Author(s):  
C. Donner ◽  
M. Gill
Keyword(s):  
Carboxylic Acid ◽  
Total Synthesis ◽  
Ochratoxin A ◽  
Propylene Oxide ◽  
Biologically Active ◽  
Acid Component ◽  
Ochratoxin Α ◽  
First Time

(R)-Ochratoxin α, the monochiral carboxylic acid component of the biologically active dipeptide ochratoxin A, is synthesized for the first time over nine steps from (R)-propylene oxide. The method constitutes a versatile and general route to functionalized dihydroisocoumarins.

scholarly journals Heterologous Expression and Characterization of A Novel Ochratoxin A Degrading Enzyme, N-acyl-L-amino Acid Amidohydrolase, from Alcaligenes faecalis

Toxins ◽  
2019 ◽  
Vol 11 (9) ◽  
pp. 518
Author(s):  
Honghai Zhang ◽  
Yunpeng Zhang ◽  
Tie Yin ◽  
Jing Wang ◽  
Xiaolin Zhang
Keyword(s):  
Amino Acid ◽  
Ochratoxin A ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Alcaligenes Faecalis ◽  
Degrading Enzyme ◽  
Sds Page ◽  
Ochratoxin Α ◽  
Acid Amidohydrolase

Ochratoxin A (OTA) is a well-known, natural contaminant in foods and feeds because of its toxic effects, such as nephrotoxicity in various animals. Recent studies have revealed that Alcaligenes faecalis could generate enzymes to efficiently degrade OTA to ochratoxin α (OTα) in vitro. In an effort to obtain the OTA degrading mechanism, we purified and identified a novel degrading enzyme, N-acyl-L-amino acid amidohydrolase (AfOTase), from A. faecalis DSM 16503 via mass spectrometry. The same gene of the enzyme was also encountered in other A. faecalis strains. AfOTase belongs to peptidase family M20 and contains metal ions at the active site. In this study, recombination AfOTase was expressed and characterized in Escherichia coli. The molecular mass of recombinant rAfOTase was approximately 47.0 kDa, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The enzyme exhibited a wide temperature range (30–70 °C) and pH adaptation (4.5–9.0) and the optimal temperature and pH were 50 °C and 6.5, respectively.

scholarly journals Analysis of the Carry-Over of Ochratoxin A from Feed to Milk, Blood, Urine, and Different Tissues of Dairy Cows Based on the Establishment of a Reliable LC-MS/MS Method

Molecules ◽  
2019 ◽  
Vol 24 (15) ◽  
pp. 2823 ◽  
Author(s):  
Zhiqi Zhang ◽  
Zhichen Fan ◽  
Dongxia Nie ◽  
Zhihui Zhao ◽  
Zheng Han
Keyword(s):  
Dairy Cows ◽  
Ochratoxin A ◽  
Limit Of Quantification ◽  
Fresh Milk ◽  
Liquid Chromatography Tandem Mass ◽  
Ochratoxin Α ◽  
Carry Over ◽  
First Time ◽  
Different Tissues

A rapid and reliable liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed for simultaneous determination of ochratoxin A (OTA) and its metabolite ochratoxin α (OTα), for the first time, in dairy cow plasma, milk, urine, heart, liver, spleen, lung, and kidney. The established method was extensively validated by determining the linearity (R2 ≥ 0.990), sensitivity (lower limit of quantification, 0.1–0.2 ng mL−1), recovery (75.3–114.1%), precision (RSD ≤ 13.6%), and stability (≥83.0%). Based on the methodological advances, the carry-over of OTA was subsequently studied after oral administration of 30 μg/kg body weight OTA to dairy cows. As revealed, OTA and OTα were detected in urine, with maximal concentrations of 1.8 ng mL−1 and 324.6 ng mL−1, respectively, but not in milk, plasma, or different tissues, verifying the protection effects of rumen flora against OTA exposure for dairy cows. Moreover, 100 fresh milk samples randomly collected from different supermarkets in Shanghai were also analyzed, and no positive samples were found, further proving the correctness of the in vivo biotransformation results. Thus, from the currently available data, regarding OTA contamination issues on dairy cows, no significant health risks were related to OTA exposure due to the consumption of these products.

A Spectrophotometric Procedure, Using Carboxypeptidase A, for the Quantitative Measurement of Ochratoxin A

1976 ◽  
Vol 59 (1) ◽  
pp. 128-129
Author(s):  
Karl Hult ◽  
Sten Gatenbeck
Keyword(s):  
Quantitative Determination ◽  
Ochratoxin A ◽  
Quantitative Measurement ◽  
Carboxypeptidase A ◽  
Excitation Spectra ◽  
Fluorescence Excitation ◽  
Fluorescence Excitation Spectra ◽  
Ochratoxin Α ◽  
The Difference

Abstract In the method described, ochratoxin A is cleaved into ochratoxin α (free isocoumarin chromophore) and phenylalanine, using carboxypeptidase. Detection is based on the difference in fluorescence excitation spectra of ochratoxin A (380 nm, maximum) and ochratoxin α (340 nm, maximum). The quantitation of ochratoxin A is based on the loss of fluorescence intensity at 380 nm. The method has been used for the quantitative determination of as little as 4 μg ochratoxin A/kg barley and barley meal but it could be extended to other products.

scholarly journals New Insight into the Ochratoxin A Biosynthetic Pathway through Deletion of a Nonribosomal Peptide Synthetase Gene in Aspergillus carbonarius

2012 ◽  
Vol 78 (23) ◽  
pp. 8208-8218 ◽  
Author(s):  
Antonia Gallo ◽  
Kenneth S. Bruno ◽  
Michele Solfrizzo ◽  
Giancarlo Perrone ◽  
Giuseppina Mulè ◽  
...  
Keyword(s):  
Ochratoxin A ◽  
Biosynthetic Pathway ◽  
Nonribosomal Peptide Synthetase ◽  
Gene Encoding ◽  
Nonribosomal Peptide ◽  
Content Type ◽  
Peptide Synthetase ◽  
Ochratoxin Α ◽  
Hydrolysis Of ◽  
Insight Into

ABSTRACTOchratoxin A (OTA), a mycotoxin produced byAspergillusandPenicilliumspecies, is composed of a dihydroisocoumarin ring linked to phenylalanine, and its biosynthetic pathway has not yet been completely elucidated. Most of the knowledge regarding the genetic and enzymatic aspects of OTA biosynthesis has been elucidated inPenicilliumspecies. InAspergillusspecies, onlypksgenes involved in the initial steps of the pathway have been partially characterized. In our study, the inactivation of a gene encoding a nonribosomal peptide synthetase (NRPS) in OTA-producingA. carbonariusITEM 5010 has eliminated the ability of this fungus to produce OTA. This is the first report on the involvement of annrpsgene product in OTA biosynthetic pathway in anAspergillusspecies. The absence of OTA and ochratoxin α, the isocoumaric derivative of OTA, and the concomitant increase of ochratoxin β, the dechloro analog of ochratoxin α, were observed in the liquid culture of transformed strain. The data provide the first evidence that the enzymatic step adding phenylalanine to polyketide dihydroisocoumarin precedes the chlorination step to form OTA inA. carbonariusand that ochratoxin α is a product of hydrolysis of OTA, giving an interesting new insight into the biosynthetic pathway of the toxin.

Removal of ochratoxin A by a carboxypeptidase and peptides present in liquid cultures of Bacillus subtilis CW14

2018 ◽  
Vol 11 (4) ◽  
pp. 559-570 ◽  
Author(s):  
H.N. Hu ◽  
X. Jia ◽  
Y.P. Wang ◽  
Z.H. Liang
Keyword(s):  
Bacillus Subtilis ◽  
Ochratoxin A ◽  
Culture Supernatant ◽  
Physical Adsorption ◽  
Proteinase K ◽  
Small Peptides ◽  
Liquid Cultures ◽  
Ochratoxin Α ◽  
Peptide Complexes

Ochratoxin A (OTA) is an important mycotoxin that contaminates a variety of agricultural products. The cell-free supernatant of Bacillus subtilis CW14 liquid cultures were reported previously to be capable of removing OTA efficiently. In this work, we examined several substances that are probably involved in this removal of OTA using in vitro experiments. The strain CW14 culture supernatant that was separated by ultrafiltration showed that the fractions collected at >10 kDa and <3 kDa had a significant ability to reduce OTA (84.9 and 74.8%, respectively) when incubated with 6 μg/ml OTA at 37 °C for 24 h. A putative metalloenzyme was responsible for the activity of the >10-kDa fraction, which was confirmed by the detrimental effects of heat treatments or addition of SDS, proteinase K, or EDTA. Subsequently, a carboxypeptidase (CP) gene that was likely related to the enzymatic conversion of OTA by the >10-kDa fraction was cloned from the B. subtilis CW14 genome, and over-expressed in Escherichia coli. The recombinant CP degraded 71.3% of OTA at 37 °C for 24 h, and ochratoxin α (OTα) was confirmed as a degradation product. From the <3-kDa fraction, some small peptides (1.7 kDa >Mw >0.7 kDa) were purified and decreased OTA by 45.0% under the same conditions, but no product was detected. These peptides were presumed to be capable of binding OTA due to their affinity with the OTA molecule, and the OTA-peptide complexes escaped from the extraction procedures for OTA quantification. These results indicated there was a probable synergistic effect that was involved in removal of OTA by the strain CW14 culture supernatant, which included enzymatic degradation by a CP and physical adsorption by some small peptides.

A superefficient ochratoxin A hydrolase with promising potential for industrial applications

2021 ◽  
Author(s):  
Han Luo ◽  
Gan wang ◽  
Nan Chen ◽  
Zemin Fang ◽  
Yazhong Xiao ◽  
...  
Keyword(s):  
Ochratoxin A ◽  
Degradation Mechanism ◽  
Kinetic Constant ◽  
Industrial Applications ◽  
Detoxification Enzymes ◽  
Toxicological Effects ◽  
Catalytic Sites ◽  
Ochratoxin Α ◽  
Detoxifying Enzyme ◽  
Temperature Adaptability

As the most seriously controlled mycotoxin produced by Aspergillus spp. and Penicillium spp., ochratoxin A (OTA) results in various toxicological effects and widely contaminates agro-products. Biological detoxification of OTA is the most priority in food and feed industry, but currently available detoxification enzymes are relatively low effectiveness in time and cost. Here we show a superefficient enzyme ADH3 identified from Stenotrophomonas acidaminiphila with a strong ability to transform OTA into non-toxic ochratoxin-α by acting as an amidohydrolase. Recombinant ADH3 (1.2 μg/mL) completely degrades 50 μg/L OTA within 90 seconds, while the availably most efficient OTA hydrolases takes several hours. The kinetic constant showed that rADH3 ( Kcat/Km ) catalytic efficiency was 56.7-35000 times higher than those of previous hydrolases rAfOTase, rOTase and commercial carboxypeptidase A (CPA). Protein structure-based assay suggested that ADH3 has a preference for hydrophobic residues to form a larger hydrophobic area than other detoxifying enzymes at the cavity of the catalytic sites, and this structure makes the OTA easier to access to catalytic sites. In addition, ADH3 shows considerable temperature adaptability to exert hydrolytic function at the temperature down to 0°C or up to 70°C. Collectively, we report a superefficient OTA detoxifying enzyme with promising potential for industrial applications. IMPORTANCE Ochratoxin A (OTA) can result in various toxicological effects and widely contaminates agro-products and feedstuffs. OTA detoxifications by microbial strains and bio-enzymes are significant to food safety. Although previous studies showed OTA could be transformed through several pathways, the ochratoxin-α pathway is recognized as the most effective one. However, the most currently available enzymes are not efficient enough. Here, a superefficient hydrolase ADH3 which can completely transform 50 μg/L OTA into ochratoxin-α within 90 seconds was screened and characterized. The hydrolase ADH3 shows considerable temperature adaptability (0-70°C) to exert the hydrolytic function. Findings of this study supplied an efficient OTA detoxifying enzyme and predicted the superefficient degradation mechanism which lay a foundation for future industrial applications.

Degradation of ochratoxin A by supernatant and ochratoxinase of Aspergillus niger W-35 isolated from cereals

2020 ◽  
Vol 13 (2) ◽  
pp. 287-298
Author(s):  
M. Zhao ◽  
X.Y. Wang ◽  
S.H. Xu ◽  
G.Q. Yuan ◽  
X.J. Shi ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Food Safety ◽  
Aspergillus Niger ◽  
Ochratoxin A ◽  
Promising Strategy ◽  
Ochratoxin Α ◽  
Food And Feed ◽  
Commercial Feeds ◽  
Penicillium Spp

Ochratoxin A (OTA) is a mycotoxin produced by Aspergillus spp. and Penicillium spp. and poses a threat to food safety. Biodegradation may be a promising strategy for reducing the OTA contamination in the future. In this study, Aspergillus niger strain W-35 was isolated from cereals and studied for its ability to degrade OTA. Results showed that the supernatant of W-35 could degrade OTA both in vitro and in commercial feeds after incubation at 37 °C for 12 h by 78.0 and 37.0%, respectively. Ochratoxin α (OTα) was assayed as a degradation product by HPLC-FLD. Furthermore, an enzyme specific for OTA degradation (ochratoxinase, OTase) obtained from W-35 was successfully expressed in Escherichia coli BL21, and degraded OTA at a rate of 85.1% for 12 h. These results indicated that this OTA degradation is enzymatic and that the responsible enzyme is extracellular OTase. Reliable degradation of OTA has the potential for wide-ranging applications in the food and feed industries.

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