Patulin

Natural Occurrence

Patulin is a polyketide lactone, produced by certain fungal species of Penicillium, Aspergillus and Byssochlamys growing on fruit, including apples, pears, grapes and other fruit. In whole fruits, visual inspection will usually identify poor-quality items. The principal risk arises when unsound fruit is used for the production of juices and other processed products. It has also been reported in vegetables, cereal grains and silage. Penicillium expansum appears to be the mould usually responsible for patulin in apple juice.

Patulin

Chemical and Physical Properties

Patulin can be isolated as colourless to white crystals from ethereal abstracts that have no optical activity. It melts at about 110 ºC and sublimes in high vacuum at 70-100 ºC. It is soluble in water, methanol, ethanol, acetone, and ethyl or amyl acetate and less soluble in diethyl ether and benzene. It undergoes all the reactions expected of a secondary alcohol, reduces warm Fehlings solution and decolorises potassium permanganate. It is stable in acid solutions but can be decomposed by boiling in 2N H2SO4 for 6 hours. It is susceptible to alkaline hydrolysis, reduced by SO2 and by fermentation.

Toxicity and Importance

Patulin possesses wide-spectrum antibiotic properties and has been tested in humans to evaluate its ability to treat common colds. However, its effectiveness has never been proven and, in the light of its toxicity, use for treatment of medical conditions has not been pursued because of its being irritant to the stomach and causing nausea and vomiting. In acute and short-term studies, patulin causes gastrointestinal hyperaemia, distension haemorrhage and ulceration.

For patulin, the LD50 for the rat has been reported as 15 mg kg-1 body and 25 mg kg-1after sub-cutaneous injection. Death was usually caused by pulmonary oedema. Lungs were oedematous, with the alveoli filled with protein-rich fluid and many leucocytes. The pulmonary vessels were congested but haemorrhages were few. Hepatic and intestinal blood vessels were congested and in sections of the kidneys there was slight congestion, mild degeneration of the tubular epithelium, and a few loci of haemorrhages. Patulin injected in large amounts over a 2-month period was carcinogenic, resulting in induction of sarcomas at the injection site. In long-term studies at lower dose levels, these effects were not observed. It has also been shown to be immunotoxic, and neurotoxic. IARC (1986) concluded that no evaluation could be made of the carcinogenicity of patulin to humans and that there was inadequate evidence in experimental animals. Based on reproduction, and long-term carcinogenicity studies in rats and mice, the JECFA allocated a Provisional Tolerable Weekly Intake of 7 µg/kg b.w.

Products Affected and Natural Occurrence

By far the most important source of patulin for humans is apples and apple juice, particularly that produced by direct pressing of apples. Other products containing apples, such as pies and jam, may contain smaller amounts. Pears, grapes, bilberries and other fruit may also be affected. Sweet cider may also be affected but only if additional apple juice is added to the fermented cider. Patulin has also been reported in vegetables, cereal grains and animal silage. Patulin is reported to be formed when silage spoils and is sometimes suspected of causing haemorrhagic symptoms in cattle, although when these incidents are investigated patulin is rarely proven to be the cause.

Sampling and Analysis

Patulin can be detected using TLC, but the usual method of analysis is by HPLC with UV detection. Ethyl acetate is used to extract patulin from aqueous-based products such as fruit juices. In some instances, solutions, particularly those of cloudy juices, require depectinisation before extraction. Other methods use gas chromatography or GC/MS, and this latter method is useful for confirmation. On-line monitoring of fruit juices for patulin would be a major benefit for the apple juice industry. In theory, suitable selectivity and sensitivity for developing such a device should be achieved by raising suitable antibodies to this mycotoxin. Incorporation of these antibodies into a monitoring device should then make it possible to produce a dip-stick test or alternatively could be used in an immunoaffinity column to assist with analytical clean-up. However, the small size of the patulin molecule has hindered the raising of suitable antibodies to date and rapid tests based on immunological principles are still awaited.

Stability and Persistence

When SO2 is used as a food preservative in fruit juice or other foods, patulin is broken down. It is not usually found in alcoholic beverages or vinegar but has been found in 'sweet' cider in which unfermented apple juice is added to the cider. Studies show that it interacts with the yeast Saccharomyces cerevisiae during fermentation and is destroyed. However, breakdown products such as ascadiol are produced and the toxicity of these is not fully understood.

In acid conditions, it is relatively stable to heat processes up to about 100 ºC. Chloroform is preferred for storage of patulin solutions as it tends to decompose in distilled water.

Legislation and Control

Up to the present, patulin has not been subject to statutory regulation in most countries. However, the quality of fruit juice is controlled in some countries by the setting of a 'guideline' or a 'recommended' maximum concentration agreed with the apple processing industry. This is commonly set at 50 µg/litre. In the UK regular monitoring of apple juice has been carried out since 1992, and this has reduced the concentrations and incidence of patulin in juices to a very low level, chiefly through better storage of apples, the avoidance of damaged or poor-quality fruit, and better production protocols.

At the 32nd session of the Codex Committee on Food Additives and Contaminants, held in March 2000, discussions took place and a limit of 50 mg/kg for patulin in apple juice and apple juice ingredients in other beverages was proposed for adoption at Step 8 to the Codex Alimentarius Commission (July 2001). In this latter meeting, a consensus could not be reached and the Commission returned the draft maximum level to Step 6 for further consideration.

Although various exposure assessments indicate that the average lifetime exposure of patulin is well below the provisional maximum tolerable daily intake (PMTDI), recent assessments indicate that the exposure of children to patulin through the consumption of apple juice is in the range of, or even exceeds, the PMDTI during a considerable period of the childhood. The Committee was of the opinion that this aspect needed a closer examination with regard to the potential health risks for children and that it was premature to adopt the level of 50 mg/kg as a maximum level for patulin in apple juice.