Patulin biodegradation by marine yeast Kodameae ohmeri

Patulin contamination of fruit- and vegetable-based products had become a major challenge for the food industry. Biological methods of patulin control can play an important role due to their safety and high efficiency. In this study, a strain of marine yeast with high patulin degradation ability was screened. The yeast was identified as Kodameae ohmeri by the BioLog identification system and partial 26S rRNA gene sequencing. The degradation products of patulin were identified as (E)- and (Z)-ascladiol through HPLC and LC-TOF/MS. High patulin tolerance at 100 μg ml^–1 and a high degradation rate at 35°C at a pH between 3 and 6 indicates the potential application of K. ohmeri for patulin detoxification of apple-derived products.     

Kinetics of the Thermal Degradation of Patulin in the Presence of Ascorbic Acid

Degradation of the mycotoxin patulin between 25 and 85 °C without and with added ascorbic acid was studied, and the effectiveness of linear and nonlinear models for predicting reaction rates was compared. In agreement with previous reports, ascorbic acid significantly increased (P ≤ 0.05) the rate of patulin degradation at all temperatures studied. The data for patulin degradation in the absence of ascorbic acid were adequately modeled using a zero‐order linear kinetic model. However, the predictive abilities of zero and higher‐order linear models were not adequate to describe the more complex reactions that likely occurred when ascorbic acid was added. In contrast, the nonlinear Weibull model adequately described the patulin‐ascorbic acid reaction throughout the temperature range studied. Zero‐order rate constants and Weibull scale values for each of the respective reactions followed the Arrhenius law. Activation energies of 58.7 ± 3.9 and 29.6 ± 1.9 kJ mol^?1 for the reaction without and with ascorbic acid, respectively, confirmed decreased patulin stability in the presence of ascorbic acid and suggested that the mechanisms for the 2 degradation reactions were different.     

The microbiology of apples and apple products

The apple industry has reached an annual production level of 8.5 billion pounds. CA storage of 25% of this crop has enabled a fresh market on a year‐round basis. To achieve high quality in raw fruit and processed apple products, careful attention must be paid to maintaining a microbiologically stable environment. The ecology of the microflora associated with the apple is a reflection of the orchard, handling, harvesting, and storage practices. Yeasts#lbpredominate on orchard fruit, molds may become a storage problem, and bacteria cause spoilage, off flavors, and loss of quality in juice products. Despite the microbial problems inherent in producing of quality product, the apple industry is faced with the occurrence of patulin. Patulin, a mycotoxin produced by Penicillium and Aspergillus species, has been associated with damaged fruit. Decreased temperatures, coupled with CA storage; can deter mold growth and patulin production. Laboratory detection methods for derivations of patulin are able to detect microgram quantities. Means to eliminate patulin formed in apple products include addition of ascorbate and SO₂, extending fermentation, or charcoal filtering. However, degradation products of patulin have not been evaluated toxicologically.     

辉光放电等离子体对水溶液中棒曲霉素的降解作用

研究辉光放电等离子体在不同酸度条件下和不同催化剂存在时对水溶液中棒曲霉素的降解作用。结果表明：当直流电压为600 V,电流范围为145～155 mA时,辉光放电等离子体可在40 min内有效降解水溶液中的棒曲霉素。在pH 4.0时降解棒曲霉素效率更高,Fe2＋、H2O2对降解反应具有催化作用,可提高棒曲霉素的降解速率,而纳米ZnO却减小了棒曲霉素的降解速率。     

二氧化钛纳米管光催化降解苹果汁中展青霉素的工艺优化

为评价二氧化钛纳米管对苹果汁中展青霉素的光催化降解性能,本文进行了光催化降解展青霉素的工艺研究,首先利用水热法制备出二氧化钛纳米管,以其光催化降解展青霉素所用时间作为活性评价指标,考察了紫外灯功率、反应温度、pH及二氧化钛纳米管的投加量对展青霉素降解效果的影响;然后据此设计正交试验进行二氧化钛纳米管光催化降解展青霉素的工艺优化。结果表明:二氧化钛纳米管与二氧化钛光催化降解模拟汁中展青霉素的活性对比结果显示,二氧化钛纳米管比二氧化钛表现出更高的光催化降解展青霉素的活性;二氧化钛纳米管光催化降解展青霉素的最佳工艺为:二氧化钛纳米管的投加量为500 mg/L,紫外灯功率为21 W,反应温度为25℃;在最佳工艺条件下,当反应体系中展青霉素的初始浓度为1000 μg/L时,二氧化钛纳米管光催化降解模拟汁与苹果汁中展青霉素的活性对比结果显示,25 min内可将模拟汁中的展青霉素含量降低到检测限以下,45 min内可将苹果汁中的展青霉素含量降低到检测限以下。     

Rapid degradation of the mycotoxin patulin in man quantified by stable isotope dilution assays

The absorption and degradation of the mycotoxin patulin in man was quantified by using a recently developed stable isotope dilution assay. Application of this currently most sensitive method revealed a patulin content less than 200 ng l -1 in the blood serum of five consumers of apple juice. Likewise, no patulin was found in the serum of a volunteer, whose blood was drawn shortly after consumption of a juice containing a maximum tolerable amount of patulin. In further in vitro experiments, the degradation of patulin by reacting it with whole blood was investigated. After addition of 100 μg patulin to 9 ml blood, only 6.1% of the mycotoxin was detected after 2 min. It was concluded, therefore, that even high naturally occurring concentrations of patulin in foods are quickly degraded before reaching other tissues than the gastrointestinal tract.     

Stability of patulin in a juice-like aqueous model system in the presence of ascorbic acid

In the present study, the stability of patulin in an aqueous juice-like model system was investigated. At acidic pH, the presence of ascorbic acid reduced the stability of patulin. After 34 days, patulin was reduced to 30% of its initial concentration in the presence of ascorbic acid compared to 68–71% in samples without ascorbic acid. Conditions during storage (presence of light, oxygen and/or metal ions) influenced the stability of patulin. Furthermore, it was possible to induce degradation of patulin by either generating hydroxyl radicals or by adding the rather stable radical diphenyl-1-picrylhydrazyl (DPPH). Data from the present study indicate that patulin is decomposed by free radicals generated by oxidation of ascorbic acid to dehydroascorbic acid. Rapid oxidation of ascorbic acid in the presence of oxygen, catalysed by free metal ions, resulted in a decrease of patulin. After complete oxidation of ascorbic acid, no further patulin degradation was observed. In contrast, slow oxidation of ascorbic acid in the presence of metal-chelators induced a continuous, slow oxidation of patulin. Due to low oxygen content in the headspace of a food package, addition of ascorbic acid to products such as apple juice, prior to filling, cannot be considered as an effective decontamination strategy.     

Removing and detoxifying methods of patulin: A review

Patulin is a toxic metabolite produced by molds, which is often found in many fruits and their products. It poses a serious threat to human health due to eating the foods contaminated by patulin frequently. So it is very important for people to find an ideal method for removing or detoxifying patulin in foods. Physical, chemical, and biological methods have been widely studied to remove or degrade it. This article reviews the latest development in the removal and detoxification of patulin using physical, chemical, and biological methods, points out their disadvantages, summarizes the degradation products and their safety of patulin, and draws the degradation pathway of patulin. Presently, no any singular method is ideal in removing or detoxifying patulin in foods, and a combination of various methods may be the better choice. Meanwhile, more attention should be paid to developing the advanced detoxification equipments during exploring the detoxification methods of patulin.     

Some factors influencing patulin production by Penicillium expansum in pome fruits

BACKGROUND: The objective of our study was to examine the effects of Penicillium expansum on patulin production in relation to isolates, species and cultivar type, incidence and severity of decay. In addition, patulin production at different incubation times and its diffusion were also investigated. These factors were evaluated in pome fruits inoculated with P. expansum and kept at 20 °C for short periods of time. RESULTS: The ability of five P. expansum isolates to grow and produce patulin in inoculated Golden Delicious apples varied among the strains from below the limit of quantification to 662 µg kg^?1. Variety and species of pome fruits influenced patulin production. P. expansum isolate PE97.IT produced a higher patulin content in apples than in pears. The highest patulin production was 386 µg kg^?1 in Golden Delicious. No blue mould symptom appeared in pears inoculated with P. expansum and no patulin was detected after 3 days at 20 °C. However, patulin increased with incubation time after 6 and 8 days. No patulin was detected in healthy pear tissue but it was high in the decayed area. CONCLUSION: Since patulin production is associated primarily with infected rotten tissue, patulin control is possible by using healthy fruits, sorting damaged and rotten fruits before processing. Copyright © 2010 Society of Chemical Industry     

降解展青霉素的乳酸菌的筛选鉴定及其降解特性研究

目的:筛选鉴定具有降解展青霉素作用的乳酸菌,并揭示其降解展青霉素的特性。方法:将从青海泡菜中分离得到的2株乳酸菌活化后与展青霉素共培养,利用高效液相色谱(HPLC)法检测共培养过程中体系中展青霉素的含量,筛选出对展青霉素具有较高降解能力的一株乳酸菌。通过对该乳酸菌形态学观察及对16S rDNA序列同源比对,确定该乳酸菌的分类进化地位。进一步准备该乳酸菌的活菌、胞内物质、胞外代谢物以及细胞壁并分别与展青霉素共培养,利用HPLC研究上述各组分对展青霉素的降解作用,探究该乳酸菌降解展青霉素的特性。结果:本研究从2株青海泡菜分离得到的乳酸菌中筛选到一株乳酸菌YZU02能够高效降解展青霉素,经微生物形态学观察及16S rDNA测序比对表明YZU02菌株为植物乳杆菌(Lactobacillus plantarum)。YZU02活菌对展青霉素降解效果较好,可在36 h内将展青霉素由10.87 μg/mL降至2.94 μg/mL。YZU02胞内物质、经展青霉素刺激后的YZU02胞内物质、YZU02胞外代谢物对展青霉素几乎没有降解作用,推测该乳酸菌去除展青霉素与生物降解无关。YZU02细胞壁可将展青霉素含量由10.84 μg/mL降低至5.10 μg/mL。结论:植物乳杆菌YZU02可通过其细胞壁的吸附作用去除溶液状态中的展青霉素,在36 h内对10.87 μg/mL的展青霉素的清除效率达到72.95%,在防治展青霉素污染方面极具应用价值。     

拟粉红锁掷孢酵母降解展青霉素的机制

展青霉素(PAT)是一类主要由曲霉属和青霉属等真菌产生的有毒次级代谢产物,主要污染水果及其制品,严重威胁着人类的健康。为了寻找一种安全高效的PAT脱毒方法,近年来采用生物法清除PAT已成为主要的研究方向。作者研究了拟粉红锁掷孢酵母(Sporidiobolus pararoseus)对PAT降解的影响及其降解机制。结果显示,S.pararoseus可以显著降低苹果伤口处PAT的积累量,体外实验发现S.pararoseus与PAT(5μg/mL)共同培养18 h,PAT可以被S.pararoseus完全降解。S.pararoseus对PAT的降解作用既不是酵母细胞壁的吸附作用,也不是细胞的吸收作用,而是细胞正常代谢下产生的胞内酶对PAT具有降解作用。本研究有助于了解S.pararoseus降解PAT的作用机制,为真菌毒素的生物降解提供了新的理论依据。     

Secondary metabolism in Penicillium expansum: Emphasis on recent advances in patulin research

The plant pathogenic fungus Penicillium expansum is a major concern of the global food industry due to its wide occurrence and ability to produce various mycotoxins, of which the most significant is patulin. Relatively less highlighted in the literature, in comparison with the other food-borne mycotoxins, patulin is one of the main factors in economic losses of vegetables and fruits. Otherwise, patulin is a health hazard which results in both short-term and long-term risks. This review includes knowledge on the biosynthetic mechanisms used for secondary metabolite production in P. expansum, with special emphasis on patulin biosynthesis. The abiotic factors triggering the production of patulin and the strategies developed to reduce or prevent the contamination by this mycotoxin are comprehensively discussed. The database presented in this review would be useful for the prioritization and development of future research.     

Saccharomyces cerevisiae YE‐7 reduces the risk of apple blue mold disease by inhibiting the fungal incidence and patulin biosynthesis

The yeast strain, Saccharomyces cerevisiae YE‐7, was tested to control the apple blue mold disease caused by Penicillium expansum and the accumulation of patulin mycotoxin. Compared with control, pre‐treatment of pathogen‐infected apples with YE‐7 significantly reduced the incidence of blue mold diseases and patulin accumulation in rotten apple tissue by 48% and 42.6%, respectively. Interestingly, late treatment of pathogen‐infected apples with YE‐7 did not decrease patulin accumulation in the rotten tissue compared with pre‐treatment or simultaneous‐treatment. Independently, patulin amount and gene expression of isoepoxydon dehydrogenase were also measured. YE‐7 pre‐treated apples showed 75% and four times reduction in patulin accumulation and IDH gene expression, respectively. The results indicated that YE‐7 directly affected patulin biosynthesis and did not affect accumulated patulin. On the other hand, cold storage enhanced the effect of YE‐7 on apple blue mold disease incidence.

Practical applications

Postharvest blue mold decay caused by Penicillium expansum was important postharvest disease of apples. In addition, under favorable conditions, P. expansum can cause severe losses in apple fruits due to the production of patulin. In this study, a strain of S. cerevisiae (YE‐7) was assessed for its efficacy not only in controlling apple blue mold diseases, but also in reducing patulin accumulation in fruit tissue. The research will give a practical way to control the decay in a bio‐control way instead of the chemical.     

Purification of patulin from Penicillium expansum culture: high-speed counter-current chromatography (HSCCC) versus preparative high-performance liquid chromatography (prep-HPLC)

Patulin is a mycotoxin produced by species of Penicillium and Aspergillus and is toxic to a wide range of organisms, including humans and livestock. To produce large amount of pure patulin for research purposes, high-speed counter-current chromatography (HSCCC) and preparative high-performance liquid chromatography (prep-HPLC) were applied to the purification of patulin. Apple juice was inoculated with P. expansum and containing 0.5 mg patulin per ml was used as a starting material for separation. For HSCCC, a biphasic solvent system consisted of ethyl acetate–hexane–pH 4 acetic acid (7.5:2.5:10, v/v/v) was used. For prep-HPLC, the separation was carried out in a C18 reversed-phase preparative column with a mobile phase containing acetonitrile–pH 4 acetic acid (5:95, v/v). Fractions containing patulin were collected and analysed by analytical HPLC and identified by congruent retention time and ultraviolet/visible (UV–VIS) spectrum of the standard. The structure of the purified patulin was confirmed by mass spectrometry and nuclear magnetic resonance. HSCCC produced 21.9 mg of patulin from 50 ml apple juice culture whereas the prep-HPLC yielded 18.1 mg. HSCCC also produced purer patulin than the prep-HPLC (98.6 versus 96.3%) and higher recovery (86.2 versus 71.3%). In addition, the HSCCC method is advantageous for its lower cost and a simpler procedure compared with the prep-HPLC. This one-step HSCCC method can potentially provide a simple, effective and environmentally friendly tool for obtaining gram-level pure patulin for toxicology, detoxification and many other patulin-related studies.     

Survival of Penicillium expansum and Patulin Production on Stored Apples after Wash Treatments

ABSTRACT: Penicillium expansum is a widespread fungus found on apples that causes fruit decay and may lead to production of a toxic secondary metabolite, patulin. This study was undertaken to evaluate the effectiveness of several chemical sanitizers against P. expansum NRRL 2304 and to establish sanitizing wash treatments that would inhibit P. expansum growth and subsequent patulin production on Empire apples destined for cider. Wash treatments included 200 ppm NaOCl, 1% StorOx®, 0.5% potassium sorbate, 300 ppm SO₂, and 0% to 5% acetic acid. Spores of P. expansum or inoculated apple slices were dipped in sanitizing wash solution for 5 min, and mold growth and patulin production was monitored on subsequent storage. It was found that 0.5% potassium sorbate and 300 ppm SO₂ did not affect mold survival or patulin production; 1% StorOx® was effective against mold spores in solution (4 log Most Probable Number destruction of spores), but there was no significant reduction in spore count when the same solution was used to sanitize mold‐inoculated apple discs. Washing with 200 ppm NaOCl delayed growth of P. expansum on inoculated apple discs but failed to completely inhibit patulin production. Acetic acid solution (2% to 5%) was the most efficient chemical against P. expansum. A wash treatment with ≥2% acetic acid for more than 1 min is recommended to completely inhibit growth of P. expansum and subsequent patulin production on apples destined for cider.     

Development of an LC-MS/MS method for the determination of pesticides and patulin in apples

A method for the simultaneous determination of 33 pesticides or degradation products together with patulin in apples by liquid chromatography-tandem mass spectrometry (LC-MS/MS) was developed. The method involved homogenization of the apples, extraction with ammonium acetate–acetic acid solution in methanol–water by ultrasonication, filtration, and determination by LC-MS/MS. The repeatability and within-laboratory reproducibility for the three spiking levels 0.02, 0.04 and 0.2 mg kg^?1 were between 4% and 35%. In general, the repeatability and reproducibility were about 10–20%. The limits of quantification (LOQs) were between 0.01 and 0.14 mg kg^?1. The method was used on incurred samples from parts of the ISAFRUIT project financed by the European Commission under the 6th Framework Programme. Samples were analysed at four different stages: after harvest, after storage (controlled), after a water bath, and after 28 days at room temperature. Pesticide residues were found at all stages, but no significant differences in the concentration were seen between the stages analysed. The concentration decreased significantly only for tolylfluanid after storage at room temperature for 28 days when only 0–6% of the original amount of tolylfluanid remained in the apples. No patulin was found in the apples stored for 28 days at room temperature and no growth of Penicillium expansum was observed on these apples. However, when the apples were inoculated with a spore suspension of P. expansum, high concentrations of patulin were found.     

Fate of patulin in the presence of the yeast Saccharomyces cerevisiae

Patulin is known to become analytically non-detectable during the production of cider from contaminated apple juice. The fate of [¹⁴C]-labelled patulin during the alcoholic fermentation of apple juice was studied. Three commercial cider strains of Saccharomyces cerevisiae degraded patulin during active fermentative growth, but not when growing aerobically. The products of patulin degradation were more polar than patulin itself and remained in the clarified fermented cider. Patulin did not appear to bind to yeast cells or apple juice sediment in these model experiments. HPLC analysis of patulin-spiked fermentations showed the appearance of two major metabolites, one of which corresponded by both TLC and HPLC to E-ascladiol prepared by the chemical reduction of patulin using sodium borohydride. Using a diode array detector, both metabolites had a λ_max = 271nm, identical to that of ascladiol. Thenmr spectrum of a crude preparation of these metabolites showed signals corresponding to those of the E-ascladiol prepared chemically and a weaker set of signals corresponding to those reported in the literature for Z-ascladiol.     

Effects of apple and pear varieties and pH on patulin accumulation by Penicillium expansum

BACKGROUND: Accumulation of patulin in apples and pears and subsequently in their by‐products is caused mostly by Penicillium expansum which causes blue mould. Fruit pH and other parameters are sometimes characteristic of a certain variety and thus the use of particular varieties might affect patulin content in the final products. The aim of this study was to evaluate the influence of fruit variety and pH on patulin accumulation. Patulin accumulation in both apple and pear juices at different pH and also in different apple varieties (Golden, Gala and Fuji) and pear varieties (Blanquilla and Conference) was assessed. RESULTS: The pH of juices significantly influenced patulin accumulation, especially in apple juice in which the highest amounts of patulin were detected at pH 3.5. In fruits, the pH values of the substrate were determinant only under cold storage. Thus, Golden apples, which presented a lower pH, accumulated more patulin at 1 °C. However, this trend was not observed at other temperatures in which varieties with higher amounts of organic acids (Golden and Fuji apples) accumulated most patulin. In the pear varieties, significant differences in pH did not lead to significant differences in patulin accumulation. CONCLUSION: Although pH influences patulin accumulation, other factors such as organic acid content may play an important role. Concerning the apple and pear varieties we studied, it seems that rather than variety, other parameters like pH or acidic content may be more important. These parameters vary significantly depending on the degree of ripeness of the fruit. Gala apple, which is used in Spanish juice production, was the only variety that accumulated dramatically higher amounts of patulin. Copyright © 2008 Society of Chemical Industry     

食品中展青霉素脱除方法研究进展

展青霉素由多种真菌产生,毒性极强,广泛存在于水果及其制品中,并通过食物链在人体富集,严重危害人类健康。本文综述了近年来物理、化学和生物方法去除和降解食品中展青霉素的研究进展,并对降解产物及其安全性进行了探讨。     

Patulin production by Paecilomyces species isolated from silage in the United Kingdom

Twenty-six Paecilomyces spp. isolated from silage in the UK were tested in a potato dextrose broth for the ability to produce the mycotoxin patulin. Of the 26 isolates 21 were shown to be capable of producing patulin. The ability of such isolates to produce patulin does not seem to be influenced by the grass species ensiled nor the pH of the silage. Patulin was not detected in any of the silage samples.