Rapid Detection and Quantification of Patulin and Citrinin Contamination in Fruits.

Patulin (PAT) and citrinin (CTN) are the most common mycotoxins produced by Penicillium and Aspergillus species and are often associated with fruits and fruit by-products. Hence, simple and reliable methods for monitoring these toxins in foodstuffs are required for regular quality assessment. In this study, we aimed to establish a cost-effective method for detection and quantification of PAT and CTN in pome fruits, such as apples and pears, using high-performance liquid chromatography (HPLC) coupled with spectroscopic detectors without the need for any clean-up steps. The method showed good performance in the analysis of these mycotoxins in apple and pear fruit samples with recovery ranges of 55-97% for PAT and 84-101% for CTN, respectively. The limits of detection (LOD) of PAT and CTN in fruits were 0.006 µg/g and 0.001 µg/g, while their limits of quantification (LOQ) were 0.018 µg/g and 0.003 µg/g, respectively. The present findings indicate that the newly developed HPLC method provides rapid and accurate detection of PAT and CTN in fruits.

Probabilistic risk assessment of patulin in imported apple juice and apple-containing beverages in Taiwan.

BACKGROUND: In Taiwan, nearly 90% of apples and raw ingredients for apple-based products are imported. Apple juices (137 samples) and apple-containing beverages (35 samples) were collected in Taiwan from imported food and analyzed for patulin content. RESULTS: The results revealed that the incidence levels of patulin for apple juice and apple-containing beverages are 5.84% and 5.71%. The mean contamination levels were 1.7 and 1.4 µg kg-1 for apple juice and apple-containing beverages. One sample exceeded 50 µg kg-1 , the maximum permissible limit according to the Taiwan Sanitation Standard for the Tolerance of Mycotoxins in Foods. The estimated intake of patulin for consumers ranged from 5.4 to 18.0 ng kg-1 body weight per day for apple juice and between 6.1 and 11.2 ng kg-1 body weight for apple-containing beverages. The highest average patulin intake was observed among infants aged 0-3 years, followed by children aged 4-12 years old. Finally, the hazard index (HI) for the 50th, 90th, and 95th percentiles are 0.0186, 0.1201, and 0.2048, respectively, for infants aged 0-3 years. CONCLUSION: The dietary intake of patulin from imported apple juices and apple-containing beverages is well below the safety levels and does not present a risk for adult and children consumers. However, it is important to point out that we only analyzed imported apple juices and apple-containing beverages. More studies are necessary to establish if the HI for patulin will still remain below 1 when other foods and beverages are included in the risk calculations. © 2020 Society of Chemical Industry.

Patulin in fruit juices: occurrence, bioaccessibility, and risk assessment for Serbian population.

This paper reports the first study of patulin occurrence in fruit juices and risk assessment related to patulin intake by children, adolescents and adults in Serbia. During 3 consecutive years (2013-2015), 142 fruit (apple or multi-fruit) juices were collected from the market and analysed using HPLC-UV. Patulin was found in 51.4% of juices with 0.7% of the samples in excess of the legal limit of 50 µg kg-1 (mean 4.3 µg kg-1). Apple juices showed significantly higher percentage of contaminated samples (74.0% vs 27.5%), as well as higher mean patulin content (6.4 vs 2.1 µg kg-1) when compared with the multi-fruit ones. Bioaccessibility of patulin in fruit juices was studied using the standardised in vitro digestion method. A mean of 21.6% of the initial patulin amount reached the end of the intestinal phase showing a significant reduction of this toxin during the human digestion process. Risk assessment of patulin intake by Serbian children, adolescents and adults, conducted by deterministic and probabilistic approaches and including the bioaccessibility results, revealed no health concern. Although patulin alone does not represent risk, further research should consider its co-occurrence with other toxic substances in food and potential adverse effects of their mixtures.

Mycoflora assessment, growth and toxigenic features of patulin-producers in kiwifruit in China.

BACKGROUND: Fungal development in agricultural products may cause mycotoxin contamination, which is a significant threat to food safety. Patulin (PAT) and PAT-producer contamination has been established as a worldwide problem. The present study aimed to investigate the mycoflora and PAT-producers present in kiwifruits and environmental samples collected from orchards and processing plants in Shaanxi Province, China. RESULTS: Variations in mycoflora were observed in different samples, with penicillia and aspergilli as the predominant genera. Approximately 42.86% of dropped fruits were contaminated with PAT-producers, which harbored the 6-methylsalicylic acid synthase and the isoepoxydon dehydrogenase genes that are involved in PAT biosynthesis. The growth of Penicillium expansum, Penicillium griseofulvum and Penicillium paneum in kiwi puree agar (KPA) medium and kiwi juice well fitted the modified Gompertz and Baranyi and Roberts models (R2 ≥ 0.95). A significant positive correlation between colony diameter and PAT content in KPA medium of P. expansum and P. griseofulvum was observed (P < 0.05). CONCLUSIONS: The present study analyzed the mycofloral composition and the potential risk for PAT and PAT-producer contamination in kiwifruit, which may be utilized in the establishment of proper management practices in the kiwifruit industry. © 2017 Society of Chemical Industry.

Risk assessment of patulin intake through apple-based food by infants and preschool children in Serbia.

This study reports for the first time in Serbia the occurrence of patulin in apple-based food and the risk assessment associated with patulin intake by infants and preschool children. In total, 214 samples of infant fruit juices (48), infant purée (66), and juices for children (small package with straw, 100) were collected over 3 years (2013-15) and analysed using HPLC with ultraviolet detection. Patulin was found in 43.8% of infant juices and 16.7% of infant purée, with all values below the legal limit of 10 µg kg-1 (maximum 8.3 and 7.7 µg kg-1, respectively). The proportion of contaminated samples among fruit juices for children was 43.0%, with the highest patulin concentration at 30.2 µg kg-1, not exceeding the maximum allowed level of 50 µg kg-1. Risk assessment of patulin intake by Serbian infants and preschool children, conducted by deterministic and probabilistic approaches, revealed a hazard quotient well below 1, indicating a tolerable exposure level and no health concern.

Patulin and ochratoxin A co-occurrence and their bioaccessibility in processed cereal-based foods: A contribution for Portuguese children risk assessment.

Patulin (PAT) and ochratoxin A (OTA) are well known enteropathogenic mycotoxins that are present in several foodstuffs. Processed cereal-based foods are among the first solid foods eaten by children, a particularly vulnerable population group. There is a lack of knowledge related to the co-occurrence of PAT and OTA in food intended for children consumption and their potential interactions during the digestion process. The present study aims to evaluate, for the first time, the co-occurrence of PAT and OTA in processed cereal-based foods for children consumption, the bioaccessibility of these two mycotoxins, and the contribution of the bioaccessibility data for human health risk assessment. PAT and OTA incidence were 75% and 50%, respectively. These mycotoxins co-occurred in 40% of analysed samples. Bioaccessibility assays revealed mean values of 52% and 56% for PAT, alone and combined with OTA; and 100% and 106% for OTA, alone and combined with PAT. Considering the human health risk assessment, and taking into account the co-occurrence and the bioaccessibility results, this study indicates a tolerable exposure to these mycotoxins representing a low risk for Portuguese children. The present work reinforces the importance of a holistic approach for risk assessment which gathers data from occurrence, exposure and bioaccessibility.

Patulin and its dietary intake by fruit juice consumption in Iran.

A survey was conducted to determine levels and dietary intake of patulin (PAT) from fruit juices consumed in Iran. PAT content was determined using high performance liquid chromatography-diode array detection in 161 samples of fruit juices including apple, pineapple, pear, peach, pomegranate and white and red grape juices. Results revealed the presence of PAT in 16.1% of the samples examined, ranging from 5 to 190.7 µg/kg. Average and median PAT content in positive samples was 34.5 and 18.5 µg/kg, respectively. PAT concentration in 2.5% of the samples was higher than the maximum limit as set by European regulations for PAT in fruit products (50 µg/kg). Estimated daily intake of PAT by fruit juice was 16.4, 45.9 and 74.6 ng/kg bw/day for Iranian adults, children and babies, respectively. The results of this study showed that PAT does not seem to be a problem in fruit-based drinks commercialised in Iran, except for apple juice.

Occurrence of patulin in organic and conventional apple-based food marketed in Catalonia and exposure assessment.

In the last years, consumption of organic foods has become increasingly popular. Nevertheless, safety of organic foods is still unclear, and needs to be thoroughly evaluated. Patulin is a mycotoxin mainly present in rotten apples and apple-based products. The aim of this study is to analyse the content of patulin in apple juices and purees derived from organic and conventional production systems, in order to assess the risk to consumers, particularly in children. A total of 93 apple-based products marketed in Catalonia were analysed, 49 of which were derived from conventional and 44 from organic farming. The results showed higher incidence of positive samples and higher concentration of patulin in organic apple purees when comparing with conventional ones. In the case of juices, significant differences were found between conventional and organic samples, but applying a multivariate analysis the type of agriculture did not seem to have a relevant contribution to patulin occurrence, being cloudiness the main factor involved. The estimated daily intake of patulin for infants and young children (0-3 years old), children (4-18 years old) and adults (19-66 years old), were below the provisional maximum tolerable daily intake (PMTDI) of 0.4 µg/kg bw in all scenarios considered.

Evaluation of strategies for reducing patulin contamination of apple juice using a farm to fork risk assessment model.

The numerous studies conducted so far on the issue of patulin contamination have focused mainly on aspects like growth of Penicillium expansum, patulin production under different conditions and the influence of processing on the patulin concentration in apple juice. The purpose of the present study was to collect the necessary information and to develop a quantitative risk assessment model (QRAM) in order to evaluate different strategies to reduce patulin contamination. For apple juice (AJ) production 3 types of apples are considered, namely fresh apples, apples stored under cold air (short term storage) and apples stored under controlled atmosphere (CA) (long term storage). The QRAM described the complete chain from the picking of apples until storage of produced AJ. In comparison to a traditional chemical analysis, the QRAM was found accurate in predicting the concentration of patulin in cloudy and clear AJs commercialised in Belgium. Simulation of the model demonstrated that the use of apples stored under CA contributes to a large extent to the patulin contamination of AJ. Since apples stored in CA are used from more or less January onwards, AJ with high patulin concentration can be produced from January onwards. It would be useful in this respect to take this into account when sampling plans are made by apple juice producers in the framework of their HACCP-system and by governments and control agencies when monitoring programmes are elaborated. The duration of deck storage between the delivery at the apple juice producer (AJP) and the processing of the apples had a large influence on the patulin concentration, and this effect was more pronounced for apples stored under controlled atmosphere compared to apples stored under cold air. The duration of the deck storage should therefore be considered as a Critical Control Point (CCP) within HACCP-systems. Also the application of a sorting step was evaluated to be efficient to reduce the high patulin concentration in AJ. Therefore, a combination of the 2 most effective measures (namely sorting out apples with an infection lesion larger than 10 cm(2) and a reduction of the volume of CA apples) was tested and resulted in a reduction to levels below 25 µg/kg in 99.7 to 99.9% of the clear and cloudy apple juices, respectively. It is therefore advisable to include a sorting step prior to processing, when apples stored in CA are used.

Assessment of dietary intake of patulin from baby foods.

Patulin is a mycotoxin produced by microscopic fungi belonging to the Penicillium and Aspergillus genera, frequently detectable in moldy fruits and their derivatives fruit products. The EC Regulation 1881/06 has imposed the limit for the presence of patulin equal to 10 µg/kg or 10 µg/L in baby food on the basis of a PMTDI of 0.4 µg/kg bw set by the Joint FAO/WHO Expert Committee on Food Additives (JECFA). A total of 120 homogenized baby foods were analyzed to evaluate the exposure of baby and children to patulin through the consumption of these products. None of examined samples has shown a toxin concentration above the limit imposed by the law, however a PAT concentration equal to 9 µg/kg was found in 22 samples, slightly below the fixed limit. The presence of patulin in marketed baby food can be regarded as a parameter indicative of the quality of raw materials used.

Variability and uncertainty assessment of patulin exposure for preschool children in Flanders.

The objective of the present study was to evaluate the patulin exposure of children consuming organic, handcrafted or conventional apple juice through a probabilistic approach and to evaluate the effectiveness of several risk management options aiming to reduce the risk for children due to patulin exposure. However, a large part of the data on patulin contamination of apple juice fell under the limit of detection (LOD). Different methods were tested to deal with these so-called left censored data and a uniform distribution with uncertain bounds was selected to handle this censorship. Variability and uncertainty assessment of patulin exposure showed that 0.9% [90% confidence interval (CI): 0.3-1.8%] of the children consuming only organic apple juice exceed the tolerable daily intake (TDI). For consumers of conventional and handcrafted apple juice this was respectively 0.1% [90% CI: 0-0.3%] and 0% [90% CI: 0-0.2%]. Reduction of the patulin contamination in apple juice to concentrations below 25 microg/kg reduced the percentage of the children exceeding the TDI to 0% [90%CI: 0-0.2%] for organic apple juice. Reduction of the apple juice consumption was less effective than a reduction of the patulin concentration in apple juice and is only useful when the patulin concentration of apple juice is below 25 microg/kg.

Occurrence of patulin in conventional and organic fruit products in Italy and subsequent exposure assessment.

The occurrence of patulin was investigated in 100 conventional and 69 organic fruity foodstuffs samples commercially available in Italy by using an HPLC method with a limit of quantification of 0.5 microg kg(-1). Patulin was detected in 26 (26%) conventional and 31 (45%) organic products with a significantly higher (p<0.01) mean concentration in the organic products (4.78 vs. 1.15 microg kg(-1)). Mean patulin concentrations in conventional apple juices, pear juices, other juices and fruits purees were 3.14, 0.22, 0.19, 0.11 microg kg(-1), respectively, and 7.11, 11.46, 2.10, 0.18 microg kg(-1) in the relevant organic products. Four samples of juices (one conventional and two organic apple, and one organic pear) contained patulin at concentrations above the limit of 50 microg kg(-1), four at concentrations between 10 and 25 microg kg(-1), and the remaining ones below 10 microg kg(-1). Patulin was detected (<1 microg kg(-1)) in only three of the 23 fruity baby food samples tested (homogenized fruits, 11 conventional and 12 organic). Based on the available data on Italian intakes of fruit juices, the estimated daily intakes of patulin, were 0.38 and 1.57 ng kg(-1) body weight (bw) from conventional and organic products, respectively. Estimated daily intakes of patulin for children were higher, 3.41 ng kg(-1) bw from conventional and 14.17 ng kg(-1) bw from organic products, but largely below the provisional maximum tolerable daily intake (PMTDI) of 400 ng kg(-1) bw. Patulin was also found in two samples of organic apple vinegar (<5 microg kg(-1)) and in fresh apples with rotten spots (12 out of 24 samples) with maximum levels at 16,402 and 44,572 microg kg(-1) for conventional and organic apples, respectively.

[Fungal spoilage of foods and its risk assessment].

From the second half of the 1990s, an increased regard has been given to the fungal spoilage of foods as follows: 1) post-harvest diseases and losses of fruits and vegetables, 2) deterioration of low water activity foods by xerophilic fungi, 3) contamination of psychrotolerant or psychrophilic fungi on foodstuffs and processed foods during storage and distribution at low temperature, and 4) spoilage of heat processed foods and soft drinks by heat-resistant fungi. In accordance with an international concern about food safety, mycotoxin contamination of foods has gained much global attention in recent times owing to its potential health hazards. The evaluation of mycotoxin hazards is principally based on the determination of a no-observed effect level (NOEL) in long-term toxicological studies, and the application of a safety factor (usually 100). In addition to hazard assessment, data on the natural occurrence of mycotoxins in various commodities and food intake data are needed to enable exposure assessment. Thus risk assessment of mycotoxins is, in fact, the product of hazard assessment and exposure assessment. In 1997, the FAO/WHO Joint Expert Committee on Food Additives (JECFA) considered estimates of the carcinogenic potency of aflatoxins and the potential risks associated with their intake. Recently the Codex Alimentarius Commission (Codex) has established standards for aflatoxin M1 in milk and for patulin in apple juice. The Codex is an international organization, supported by FAO/WHO, aiming at facilitating world trade and protecting the health of the consumer by developing international standards for food and feeds. Apart from aflatoxins, the JECFA has measured a provisional tolerable daily intake (TDI) for ochratoxin A, patulin, deoxynivalenol, T-2/HT-2 toxins, zearalenone and fumonisins. In 2001, the mycotoxins evaluated or re-evaluated at the JECFA meeting included ochratoxin A, deoxynivalenol, T-2/HT-2 toxins, fumonisins, and aflatoxin M1. In Japan, specific regulations now exist for deoxynivalenol (1.1 ppm) in wheat grains and for patulin (50 ppb) in apple juice and its products.

Patulin in domestic and imported apple-based drinks in Belgium: occurrence and exposure assessment.

Apple-based beverages are regularly consumed by adults and children in Belgium. They are locally produced or imported from other countries. The apples used as starting material for these productions are frequently contaminated by mycotoxin-producing moulds and damaged during transport and handling. The current study was undertaken to investigate whether patulin (PAT) is present in the industrial or handicraft-made apple juices and ciders consumed by the Belgian population and to assess the population's exposure to this mycotoxin through apple-based drinks. Belgian (n = 29) and imported (14) apple juices as well as ciders (7) were assayed for PAT by high-performance liquid chromatography with ultraviolet light detection. PAT was detected respectively in 79, 86 and 43% of these tested samples. However, no contaminated sample exceeded the safe level of 50 microg PAT l(-1). Levels of PAT contamination were comparable in Belgian and imported juice samples. The overall mean PAT concentrations were 9.0 and 3.4 microg l(-1) for contaminated apple juices and ciders, respectively. This study also indicates that there was no statistically significant difference in the mean PAT contamination between clear (7.8 microg l(-1)) and cloudy (10.7 microg l(-1)) apple juices, as well as between handicraft-made apple juices (14.6 microg l(-1)) and industrial ones (7.0 microg l(-1)). On the basis of the mean results, a consumer exposure assessment indicates that a daily intake of 0.2 litres apple juice contributes to 45% of the provisional maximum tolerable daily intake for a child of 10 kg body weight.

International proficiency testing of analytical laboratories for foods and feeds from 1990 to 1996: the experiences of the United Kingdom Food Analysis Performance Assessment Scheme.

The Food Analysis Performance Assessment Scheme (FAPAS) organized by a Secretariat of the UK Ministry of Agriculture, Fisheries, and Food has checked the proficiency of analytical laboratories for foods and feeds from 1990 to 1996. FAPAS was started for UK laboratories but was expanded worldwide at the request of analysts in other countries who did not have a home-based scheme. Thirteen thousand homogeneity-checked test materials were issued, covering a very wide range of analytes, including pesticides, toxins, veterinary drug residues, trace and nutritional elements, food colors, preservatives, sweeteners, alcohol congeners, fatty acids, nitrate, and proximate analysis. Participants returned 85% of requested data, and 47,000 z-score proficiency assessments were made, of which 81% were satisfactory. Evidence is presented of improvements in overall analytical ability with increased participation in proficiency testing in the areas of proximate analysis; organochlorine pesticide analysis; and lead, mercury, and acesulfame-K analyses. Little improvement was shown in other analytical areas such as calcium analysis. Overall accuracies for analysis of specific pesticides and specific trace elements in the circulated test materials were compared.