Lipophilic toxin profiles detected in farmed and benthic mussels populations from the most relevant production zones in Southern Chile

Lipophilic toxins associated with diarrhoeic toxins were found in Mytilus chilensis (Blue mussels) and Aulacomya ater (Ribbed mussels). These shellfish samples were collected from Chiloe Island, Southern Chile. The samples were tested by liquid chromatography-tandem mass spectrometry (LC-MS/MS). After the analysis, four toxins were found: DTX-1, DTX-3, YTX and PTX. All toxins were identified by comparing their HPLC retention times with those of analytical standards and confirmed by LC-MS/MS. Dinophysistoxin-1 (DTX-1) and dinophysistoxin-3 (DTX-3) toxins were the major components within the mussel extracts. Nevertheless, the percentages of these toxins differed depending on the area they were collected from and/or the sampling date. The levels detected in Butacheuques Island for okadaic acid (OA) was 267 ± 3.5 μg OA eq kg(-1) (p < 0.05) and for DTX-3 was 183.4 ± 7.5 μg kg(-1) in ribbed mussels. Pectenotoxin (PTX) and yessotoxin (YTX) were the toxins detected in minor proportions in the toxic profile of the bivalves. The maximum concentration of YTX detected in ribbed mussels was 85.2 ± 2.8 μg kg(-1) in Mechuque Island, whereas the PTX-2 level in ribbed mussels was 82.0 ± 2.4 μg kg(-1) in Cailin Island. Analogues of YTX and PTX-2 were not detected in any of the analysed mussels, which did not support the supposed presence of isomers of toxins as a result of the enzymatic metabolism of bivalves. This study found evidence proving co-occurrence of lipophilic toxins - like PTX and YTX - with diarrhoeic toxin in samples collected in Southern Chile, which is, to date, the more complex mix of lipophilic toxins ever found in mussels samples from Southern Chile.     

In vitro biotransformation of OA-group and PTX-group toxins in visceral and non-visceral tissues of Mytilus chilensis and Ameghinomya antiqua

ABSTRACT

Lipophilic marine toxins (LMTs) are made up of multiple groups of toxic analogues, which are characterised by different levels of cellular and toxic action. The most prevalent groups in the southern Pacific zone are: a) okadaic acid group (OA-group) which consists of okadaic acid (OA) and dinophysistoxin-1 (DTX-1); and, b) pectenotoxin-2 (PTX2) group which consists of pectenotoxin-2 (PTX-2). The main objective of our study was to examine in vitro biotransformation of OA-group and PTX-group in the tissues of two endemic species of bivalves from southern Chile; blue mussels (Mytilus chilensis) and clams (Ameghinomya antiqua). The biotransformation processes of both groups were only detected in the digestive glands of both species using LC-MS/MS. The most frequently detected analogues were acyl derivatives (≈2.0 ± 0.1 μg ml^?1) for OA-group and PTX-2SA (≈1.4 ± 0.1 μg ml^?1) for PTX-group, with a higher percentage of biotransformation for OA-group (p < .001). In addition, simultaneous incubations of the different analogues (OA/PTX-2; DTX-1/PTX-2 and OA/DTX-1/PTX-2) did not show any interaction between the biotransformation processes. These results show that the toxicological variability of endemic species leads to biotransformation of the profile of toxins, so that these new analogues may affect people’s health.     

Determination of the variability of both hydrophilic and lipophilic toxins in endemic wild bivalves and carnivorous gastropods from the Southern part of Chile

The aim of this study was to analyse and determine the composition of paralytic shellfish poisoning (PSP) toxins and lipophilic toxins in the Region of Aysén, Chile, in wild endemic mussels (Mytilus chilensis, Venus antiqua, Aulacomya ater, Choromytilus chorus, Tagelus dombeii and Gari solida) and in two endemic carnivorous molluscs species (Concholepas concholepas and Argobuccinum ranelliforme). PSP-toxin contents were determined by using HPLC with fluorescence detection, while lipophilic toxins were determined by using LC-MS/MS. Mean concentrations for the total of PSP toxins were in the range 55–2505 μg saxitoxin-equivalent/100 g. The two most contaminated samples for PSP toxicity were bivalve Gari solida and carnivorous Argobuccinum ranelliforme with 2505 ± 101 and 1850 ± 137 μg saxitoxin-equivalent/100 g, respectively (p < 0.05). The lipophilic toxins identified were okadaic acid, dinophysistoxin-1 (DTX-1), azaspiracid-1 (AZA-1), pectenotoxin-2 (PTX-2) and yessotoxins (YTX). All analysed molluscs contained lipophilic toxins at levels ranging from 56 ± 4.8 to 156.1 ± 8.2 μg of okadaic acid-equivalent/kg shellfish together with YTX at levels ranging from 1.0 ± 0.1 to 18 ± 0.9 μg of YTX-equivalent/kg shellfish and AZA at levels ranging from 3.6 ± 0.2 to 31 ± 2.1 μg of AZA-equivalent/kg shellfish. Furthermore, different bivalves and gastropods differ in their capacity of retention of lipophilic toxins, as shown by the determination of their respective lipophilic toxins levels. In all the evaluated species, the presence of lipophilic toxins associated with biotransformation in molluscs and carnivorous gastropods was not identified, in contrast to the identification of PSP toxins, where the profiles identified in the different species are directly related to biotransformation processes. Thus, this study provides evidence that the concentration of toxins in the food intake of the evaluated species (Bivalvia and Gastropoda class) determines the degree of bioaccumulation and biotransformation they will thereafter exhibit.     

Saxitoxins and okadaic acid group: accumulation and distribution in invertebrate marine vectors from Southern Chile

Harmful algae blooms (HABs) are the main source of marine toxins in the aquatic environment surrounding the austral fjords in Chile. Huichas Island (Aysén) has an history of HABs spanning more than 30 years, but there is limited investigation of the bioaccumulation of marine toxins in the bivalves and gastropods from the Region of Aysén. In this study, bivalves (Mytilus chilenses, Choromytilus chorus, Aulacomya ater, Gari solida, Tagelus dombeii and Venus antiqua) and carnivorous gastropods (Argobuccinum ranelliformes and Concholepas concholepas) were collected from 28 sites. Researchers analysed the accumulation of STX-group toxins using a LC with a derivatisation post column (LC-PCOX), while lipophilic toxins (OA-group, azapiracids, pectenotoxins and yessotoxins) were analysed using LC-MS/MS with electrospray ionisation (+/–) in visceral (hepatopancreas) and non-visceral tissues (mantle, adductor muscle, gills and foot). Levels of STX-group and OA-group toxins varied among individuals from the same site. Among all tissue samples, the highest concentrations of STX-group toxins were noted in the hepatopancreas in V. antiqua (95 ± 0.1 μg STX-eq 100 g^?1), T. dombeii (148 ± 1.4 μg STX-eq 100 g^?1) and G. solida (3232 ± 5.2 μg STX-eq 100 g^?1; p < 0.05); in the adductor muscle in M. chilensis (2495 ± 6.4 μg STX-eq 100 g^?1; p < 0.05) and in the foot in C. concholepas (81 ± 0.7 μg STX-eq 100 g^?1) and T. dombeii (114 ± 1.2 μg STX-eq 100 g^?1). The highest variability of toxins was detected in G. solida, where high levels of carbamate derivatives were identified (GTXs, neoSTX and STX). In addition to the detected hydrophilic toxins, OA-group toxins were detected (OA and DTX-1) with an average ratio of ≈1:1. The highest levels of OA-group toxins were in the foot of C. concholepas, with levels of 400.3 ± 3.6 μg OA eq kg^?1 (p < 0.05) and with a toxic profile composed of 90% OA. A wide range of OA-group toxins was detected in M. chilensis with a toxicity < 80 μg OA eq kg^?1, but with 74% of those toxins detected in the adductor muscle. In all evaluated species, there was no detection of lipophilic toxins associated with biotransformation in molluscs and carnivorous gastropods. In addition, the STX-group and OA-group toxin concentrations in shellfish was not associated with the presence of HAB. The ranking of toxin concentration in the tissues of most species was: digestive glands > mantle > adductor muscle for the STX-group toxins and foot > digestive gland for the OA-group toxins. These results gave a better understanding of the variability and compartmentalisation of STX-group and OA-group toxins in different bivalve and gastropod species from the south of Chile, and the analyses determined that tissues could play an important role in the biotransformation of STX-group toxins and the retention of OA-group toxins.     

Ergot alkaloids in rye flour determined by solid-phase cation-exchange and high-pressure liquid chromatography with fluorescence detection

Ergot alkaloids are mycotoxins that are undesirable contaminants of cereal products, particularly rye. A method was developed employing clean-up by cation-exchange solid-phase extraction, separation by high-performance liquid chromatography under alkaline conditions and fluorescence detection. It is capable of separating and quantifying both C8-isomers of ergocornine, α-ergocryptine, ergocristine, ergonovine, and ergotamine. The average recovery was 61% ±?10% with limits of detection from 0.2 to 1.1 µg kg^?1. Twenty-four unknown rye flour samples from Danish mills contained on average 46 µg kg^?1 with a maximum content of 234 µg kg^?1. The most common ergot alkaloids were ergotamine and α-ergocryptine including their C8-isomers. A total of 54% of the ergot alkaloids were detected as C(8)-S isomers.     

Evaluation and validation of two microbiological tests for screening antibiotic residues in honey according to the European guideline for the validation of screening methods

Two microbiological kits based on Bacillus stearothermophilus (Eclipse 50® and Premi®Test) have been evaluated and validated according to the European guideline for the validation of screening methods (January 2010) and in relation to the concentrations recommended by the EU-RL in 2007. Both tests are robust, a fast method and easy to implement. Both tests are applicable to a very large variety of honeys from different floral and geographical origins (rosemary, lavender, scrub, heath, alder, forest, lemon, acacia, chestnut, raspberry, mountain and flowers) as well as honey of different colours (from blank honey to brown honey, including yellow and orange honey). A satisfactory false-positive rate of 5% was obtained for the Eclipse 50® test. The observed detection capabilities CCβ of the Eclipse 50® kit were: chlortetracycline (>75?µg?kg^?1), oxytetracycline (≤200?µg?kg^?1), tetracycline (>100?µg?kg^?1), cloxacillin (≤40?µg?kg^?1), tylosin (≤200?µg?kg^?1), desmycosin (>400?µg?kg^?1), sulfadiazine (≤300?µg?kg^?1), sulfadimethoxine (≤250?µg?kg^?1), sulfamerazine (>300?µg?kg^?1), sulfamethazine (>1000?µg?kg^?1), sulfamethizole (>75?µg?kg^?1), sulfamethoxazole (≤25?µg?kg^?1), sulfanilamide (?1000?µg?kg^?1), sulfaquinoxaline (>75?µg?kg^?1), sulfathiazole (≤250?µg?kg^?1) and lincomycin (>1500?µg?kg^?1). These levels were all higher than the recommended concentrations where they exist. Due to its lack of sensitivity, it cannot be recommended for reliable routine use. The observed CCβ of the Premi®Test kit were: chlortetracycline (10?µg?kg^?1), oxytetracycline (>10?µg?kg^?1), tetracycline (≤10?µg?kg^?1), cloxacillin (≤5?µg?kg^?1), tylosin (≤10?µg?kg^?1), desmycosin (≤15?µg?kg^?1), sulfadiazine (≤25?µg?kg^?1), sulfadimethoxine (≤25?µg?kg^?1), sulfamerazine (≤25?µg?kg^?1), sulfamethazine (≤25?µg?kg^?1), sulfamethizole (≤25?µg?kg^?1), sulfamethoxazole (≤10?µg?kg^?1), sulfanilamide (≤25?µg?kg^?1), sulfaquinoxaline (≤10?µg?kg^?1), sulfathiazole (25?µg?kg^?1) and lincomycin (≤25?µg?kg^?1). The Premi®Test kit could be recommended for reliable use in routine control due to its low detection capabilities (except for aminoglycosides), but the disadvantage is a high false-positive rate of 14%.     

Survey of T-2 and HT-2 toxins by LC–MS/MS in oats and oat products from European oat mills in 2005–2009

T-2 and HT-2 toxins were analysed in oats (n?=?243), oat flakes (n?=?529), oat meal (n?=?105) and oat by-products (n?=?209) from 11 European mills during 2005–2009 by high-performance liquid chromatography with a triple quadrupole mass spectrometer. Limits of quantification were 5?µg?kg^?1 for both T-2 and HT-2 toxins in oats. The incidence of T-2?+?HT-2 (>5?µg?kg^?1) in oats, oat flakes, oat meal and oat by-products was 93, 77, 34 and 99%, respectively. The mean values of T-2?+?HT-2 were 94, 17, 11 and 293?µg?kg^?1 for oats, oat flakes, oat meal and oat by-products, respectively. T-2 and HT-2 occurred together and the T-2 level was 52% of HT-2 in oats. Maximal T-2 and HT-2 concentration in oat flakes and oat meal were 197 and 118?µg?kg^?1. The toxins were reduced by 82–88% during processing, but increased 3.1 times in oat by-products.     

Development and application of a non-targeted extraction method for the analysis of migrating compounds from plastic baby bottles by GC-MS

In 2011, the European Union prohibited the production of polycarbonate (PC) baby bottles due to the toxic effects of the PC monomer bisphenol-A. Therefore, baby bottles made of alternative materials, e.g. polypropylene (PP) or polyethersulphone (PES), are currently marketed. The principal aim of the study was the identification of major compounds migrating from baby bottles using a liquid–liquid extraction followed by GC/MS analysis. A 50% EtOH in water solution was selected as a simulant for milk. After sterilisation of the bottle, three migration experiments were performed during 2 h at 70°C. A non-targeted liquid–liquid extraction with ethyl acetate–n-hexane (1:1) was performed on the simulant samples. Identification of migrants from 24 baby bottles was done using commercially available WILEY and NIST mass spectra libraries. Differences in the migrating compounds and their intensities were observed between the different types of plastics, but also between the same polymer from a different producer. Differences in the migration patterns were perceived as well between the sterilisation and the migrations and within the different migrations. Silicone, Tritan? and PP exhibited a wide variety of migrating compounds, whereas PES and polyamide (PA) showed a lower amount of migrants, though sometimes in relatively large concentrations (azacyclotridecan-2-one up to 250 µg kg^?1). Alkanes (especially in PP bottles), phthalates (dibutylphthalate in one PP bottle (±40 µg kg^?1) and one silicone bottle (±25 µg kg^?1); diisobutylphthalate in one PP (±10 µg kg^?1), silicone (up to ±80 µg kg^?1); and Tritan? bottle (±30 µg kg^?1)), antioxidants (Irgafos 168, degradation products of Irganox 1010 and Irganox 1076), etc. were detected for PP, silicone and Tritan? bottles. Although the concentrations were relatively low, some compounds not authorised by European Union Regulation No. 10/2011, such as 2,4-di-tert-butylphenol (10–100 µg kg^?1) or 2-butoxyethyl acetate (about 300 µg kg^?1) were detected. Migrating chemicals were identified as confirmed (using a standard) or as tentative (further confirmation required).     

Simultaneous determination of type A,B and D trichothecenes and their occurrence in cereals and cereal products

A sensitive LC–MS/MS method for the simultaneous determination of type A, B and D trichothecenes in cereals is presented. The limits of detection ranged between 0.1 and 0.7 µg kg^?1 for all analytes. The method was applied to 289 representatively drawn samples of wheat, rye and oat products. Ninety-four percent of the wheat samples (n = 130), 95% of the rye samples (n = 61) and 100% of the oat samples (n = 98) were contaminated with the type A trichothecenes T-2 and HT-2 toxin. Median levels of T-2/HT-2 (sum of the toxins) were 0.91, 0.53 and 8.2 µg kg^?1, respectively. Highest levels were found in wheat bran (24 µg kg^?1), rye kernels (3.1 µg kg^?1) and oat flakes (85 µg kg^?1). All wheat and rye samples and 75% of the oat samples were contaminated with the type B trichothecene deoxynivalenol. Median levels of this toxin were 23, 15 and 0.53 µg kg^?1, respectively. Highest levels were found in wheat bran (1160 µg kg^?1), rye kernels (288 µg kg^?1) and oat flakes (55 µg kg^?1). The type B trichothecene nivalenol was detected in 67% of the wheat samples, in 3% of the rye samples and in 24% of the oat samples with highest levels in wheat bran (96 µg kg^?1), rye kernels (1.8 µg kg^?1) and in oat flakes (17 µg kg^?1), respectively. Levels of other type A and B trichothecenes played a minor role, although the rates of contamination were often high. Neither macrocyclic type D trichothecenes (satratoxin G and H, verrucarin A, roridin A) nor diacetylverrucarol and verrucarol (type A trichothecenes), were detected in any of the samples.     

Transfer of aflatoxin B1 and fumonisin B1 from naturally contaminated raw materials to beer during an industrial brewing process

The aim of this research was to determine the fate of aflatoxins (AFs) and fumonisins (FBs) naturally occurring in raw materials (maize grit and malted barley) during four industrial brewing processes. The aflatoxin B₁ (AFB₁) level in raw materials varied from 0.31 to 14.85 µg kg^?1, while the fumonisin B₁ (FB₁) level (only in maize grit) varied from 1146 to 3194 µg kg^?1. The concentration in finished beer ranged from 0.0015 to 0.022 µg l^?1 for AFB₁ and from 37 to 89 µg l^?1 for FB₁; the other aflatoxins and fumonisin B₂ were not found in beer samples. The average percentage of toxins recovered in finished beer, referring to the amounts contained in raw materials, were 1.5% ± 0.8% for AFB₁ and 50.7% ± 4.7% for FB₁. These results were mainly due to the different solubility of the two mycotoxins during the mashing process. If raw materials comply with the limits fixed by European Commission Regulations, the contribution of a moderate daily consumption of beer to AFB₁ and FB₁ intake does not contribute significantly to the exposure of the consumer.     

Natural occurrence of type-B trichothecene mycotoxins in Korean cereal-based products

Type-B trichothecenes (deoxynivalenol (DON), nivalenol (NIV), fusarenone-X (FUS-X), 15-acetyldeoxynivalenol (15ADON), and 3-acetyldeoxynivalenol (3ADON)) were determined in 338 cereal-based products. Detection limit, quantification limit and mean recovery for five toxins were in the ranges 0.7–2.6?µg?kg^?1, 2.1–7.8?µg?kg^?1 and 73–110%, respectively. The range of occurrence and average level in samples were, respectively, 21–88% and 5.2–121.8?µg?kg^?1 for NIV, 10–96% and 1.7–109.5?µg?kg^?1 for DON, 2–39% and 0.4–3.6?µg?kg^?1 for FUS-X, 0–80% and 0–17.3?µg?kg^?1 for 15ADON, and 0–29% and 0–1.5?µg?kg^?1 for 3ADON. Regarding co-occurrence, 64% of samples had more than two type-B trichothecenes. The estimated daily intakes of NIV, DON, FUS-X, 15ADON, and 3ADON were 0.077, 0.048, 0.004, 0.006 and 0.002?µg?kg^?1?bw?day^?1, respectively. These results suggest that current exposure levels do not indicate the possibility of adverse effects, but consideration of the combined exposure of type-B trichothecenes may be required due to the high frequency of co-occurrence.     

Occurrence and intake of deoxynivalenol in cereal-based products marketed in Korea during 2007–2008

The occurrence of deoxynivalenol (DON) was investigated in 514 cereal-based products (corn-based, n = 125; barley-based, n = 96; wheat-based, n = 94; rice-based, n = 199) marketed in Korea during 2007?2008, and estimates of DON intake were determined. Samples were analysed by high-performance liquid chromatography (HPLC) with ultraviolet light (UV) detection after immunoaffinity clean-up. The limits of detection (LOD) and limits of quantification (LOQ) were 2.2 and 5.6 µg kg^–1, respectively. Recoveries and repeatability expressed as coefficients of variation (CV) were 82.3–100% and 2.4–15.3% in beer, bread and dried corn. The incidences and mean levels of DON were 56% and 68.9 µg kg^?1 for corn-based products, 49% and 24.1 µg kg^?1 for wheat-based products, 43% and 7.5 µg kg^?1 for barley-based products, and 16% and 3.4 µg kg^?1 for rice-based products, respectively. The estimated daily intake of DON from the consumption of rice-based, wheat-based, barley-based and corn-based products were 0.0038 µg kg^?1 bw day^?1, 0.0032 µg kg^?1 bw day^?1, 0.0015 µg kg^?1 bw day^?1 and 0.0002 µg kg^?1 bw day^?1, respectively. These values represent 0.38%, 0.32%, 0.25% and 0.01% of the provisional maximum tolerable daily intake (PMTDI) of 1 µg kg^?1 bw day^?1. These results indicate that rice-based products are major contributors to DON exposure in Korea, even though the current exposure level is unlikely to cause adverse health effects.     

Fungal and bacterial metabolites in commercial poultry feed from Nigeria

Metabolites of toxigenic fungi and bacteria occur as natural contaminants (e.g. mycotoxins) in feedstuffs making them unsafe to animals. The multi-toxin profiles in 58 commercial poultry feed samples collected from 19 districts in 17 states of Nigeria were determined by LC/ESI–MS/MS with a single extraction step and no clean-up. Sixty-three (56 fungal and seven bacterial) metabolites were detected with concentrations ranging up to 10,200?µg?kg^-1 in the case of aurofusarin. Fusarium toxins were the most prevalent group of fungal metabolites, whereas valinomycin occurred in more than 50% of the samples. Twelve non-regulatory fungal and seven bacterial metabolites detected and quantified in this study have never been reported previously in naturally contaminated stored grains or finished feed. Among the regulatory toxins in poultry feed, aflatoxin concentrations in 62% of samples were above 20?µg?kg^?1, demonstrating high prevalence of unsafe levels of aflatoxins in Nigeria. Deoxynivalenol concentrations exceeded 1000?µg?kg^?1 in 10.3% of samples. Actions are required to reduce the consequences from regulatory mycotoxins and understand the risks of the single or co-occurrence of non-regulatory metabolites for the benefit of the poultry industry.     

Feed additives diclazuril and nicarbazin in egg and liver samples from Croatian farms

In total 307 egg and 275 liver samples were examined for nicarbazin and 365 eggs for diclazuril over a 30-month period. Enzyme-linked immunosorbent assay methods used for quantification were validated according to European Commission Decision 2002/657/EC. Non-compliant samples were confirmed by LC-MS/MS. Mean diclazuril concentrations in egg samples were 0.31?µg?kg^?1, which is below the MRL. In only one egg sample, 2.26?µg?kg^?1 was determined by enzyme-linked immunosorbent assay, although confirmation by LC-MS/MS gave a value of 1.6?µg?kg^?1. Mean nicarbazin levels determined were 1.85?µg?kg^?1 in egg and 21.1?µg?kg^?1 in liver samples. Four samples, one egg and three livers, yielded elevated concentrations of nicarbazin, but only in the egg sample the LC-MS/MS method confirmed nicarbazin (106?µg?kg^?1) above the MRL value.     

Isotope dilution-GC-MS/MS analysis of 16 polycyclic aromatic hydrocarbons in selected medicinal herbs used as health food additives

Medicinal herbs have a very important role in health protection and disease control, and have been used in health foods. Polycyclic aromatic hydrocarbons (PAHs) have carcinogenic, biological and mutagenic effects. In this paper, the content of 16?PAHs as representative contaminants in nine Chinese medicinal herbs, as additives for health foods, was investigated in order to ensure food safety from this source. A highly sensitive isotope dilution-gas chromatography-tandem mass spectrometry (ID-GC-MS/MS) method combined with gel permeation chromatography (GPC) and solid-phase extraction (SPE) was developed. Calibration curves showed good linearity for all PAHs (R ^2?>?0.999), and the limit of quantification (LOQ) ranged from 0.42 to 2.7?µg?kg^?1. Average recoveries for these compounds were in the range of 52.5–117%, 52.6–119% and 81.4–108% at the concentrations of 10, 50 and 250?µg?kg^?1 with RSD of 1.8–15%, 0.9–15% and 1.0–15%, respectively. The proposed method was used for the analysis of nine Chinese medicinal herbs. Total levels of PAHs varied from 98.2?µg?kg^?1 (cassia seed) to 2245?µg?kg^?1 (eucommia bark). The highest level was found for phenanthrene (Phe) in liquorice root (631.3?µg?kg^?1), indigowoad leaf (551.0?µg?kg^?1), rose flower (435.2?µg?kg^?1) and eucommia bark (432.3?µg?kg^?1). The proposed method could provide a useful basis for safety monitoring of herbs and risk management for PAHs in the health food industry.     

Pyrrolizidine alkaloids in honey: comparison of analytical methods

Pyrrolizidine alkaloids (PAs) are a structurally diverse group of toxicologically relevant secondary plant metabolites. Currently, two analytical methods are used to determine PA content in honey. To achieve reasonably high sensitivity and selectivity, mass spectrometry detection is demanded. One method is an HPLC-ESI-MS-MS approach, the other a sum parameter method utilising HRGC-EI-MS operated in the selected ion monitoring mode (SIM). To date, no fully validated or standardised method exists to measure the PA content in honey. To establish an LC-MS method, several hundred standard pollen analysis results of raw honey were analysed. Possible PA plants were identified and typical commercially available marker PA-N-oxides (PANOs). Three distinct honey sets were analysed with both methods. Set A consisted of pure Echium honey (61–80% Echium pollen). Echium is an attractive bee plant. It is quite common in all temperate zones worldwide and is one of the major reasons for PA contamination in honey. Although only echimidine/echimidine-N-oxide were available as reference for the LC-MS target approach, the results for both analytical techniques matched very well (n?=?8; PA content ranging from 311 to 520?µg?kg^?1). The second batch (B) consisted of a set of randomly picked raw honeys, mostly originating from Eupatorium spp. (0–15%), another common PA plant, usually characterised by the occurrence of lycopsamine-type PA. Again, the results showed good consistency in terms of PA-positive samples and quantification results (n?=?8; ranging from 0 to 625?µg?kg^?1 retronecine equivalents). The last set (C) was obtained by consciously placing beehives in areas with a high abundance of Jacobaea vulgaris (ragwort) from the Veluwe region (the Netherlands). J. vulgaris increasingly invades countrysides in Central Europe, especially areas with reduced farming or sites with natural restorations. Honey from two seasons (2007 and 2008) was sampled. While only trace amounts of ragwort pollen were detected (0–6.3%), in some cases extremely high PA values were detected (n?=?31; ranging from 0 to 13019?µg?kg^?1, average?=?1261 or 76?µg?kg^?1 for GC-MS and LC-MS, respectively). Here the results showed significantly different quantification results. The GC-MS sum parameter showed in average higher values (on average differing by a factor 17). The main reason for the discrepancy is most likely the incomplete coverage of the J. vulgaris PA pattern. Major J. vulgaris PAs like jacobine-type PAs or erucifoline/acetylerucifoline were not available as reference compounds for the LC-MS target approach. Based on the direct comparison, both methods are considered from various perspectives and the respective individual strengths and weaknesses for each method are presented in detail.     

Toxicity profile of commercially produced indigenous banana beer

Mycotoxins, together with endotoxins, represent important classes of naturally occurring contaminants in food products, posing significant health risks to consumers. The aim of this study is to investigate the occurrence of both Fusarium mycotoxins and endotoxins in commercially produced traditional banana beer. Two brands of commercially produced traditional banana beer were collected from a local retail market in Kigali, Rwanda. Beer samples were analysed for the presence of deoxynivalenol (DON), fumonisin B₁ and zearalenone (ZEA), using an enzyme-linked immuno-sorbent assay (ELISA) method. The quantification of bacterial endotoxin using Limulus amoeboecyte lysate (LAL) assay was also conducted. The contamination levels were 20 and 6.7?µg?kg^?1 for DON; 34 and 31.3?µg?kg^?1 for FB₁; 0.66 and 2.2?µg?kg^?1 for ZEA in brands A and B of the beers, respectively. Results indicate that the levels of Fusarium toxins and bacterial endotoxin reported in this study did not pose adverse human health effects as a result of drinking/consuming banana beer. However, exposure to low/sub-threshold doses or non-toxic levels of endotoxins magnifies the toxic effect of xenobiotic agents (e.g. fungal toxins) on liver and other target organs. Considering Fusarium toxins and/or endotoxin contamination levels in other agricultural commodities intended for human consumption, health risks might be high and the condition is aggravated when beer is contaminated by mixtures of the mycotoxins, as indicated in this study.     

Occurrence of Fusarium T-2 and HT-2 toxins in oats from cultivar studies in Germany and degradation of the toxins during grain cleaning treatment and food processing

For the Fusarium trichothecene mycotoxins T-2 and HT-2, a combined (T-2 + HT-2) temporary tolerable daily intake (tTDI) of 0.06 µg kg^?1 body weight day^?1 was proposed at the European level in 2001 (Opinion of the Scientific Committee on Food). In the near future, maximum levels for these trichothecenes will be regulated by the European Commission as announced in EU (VO) 1881/2006. For the implementation of these maximum levels, more data on occurrence and behaviour of T-2 and HT-2 toxins in primary agricultural products as well as during cleaning treatment and food processing are needed. In the current work, we determined the T-2/HT-2 concentrations in four oat cultivars (Aragon, Dominik, Ivory, Pergamon) from ten different agricultural sites in Germany, grown in cultivar studies in 2007. The grains were de-hulled, oat meal was prepared, and bread with 20% oat meal and 80% wheat flour was baked. In the cereal-processing chain, samples were taken at various steps and subsequently analysed for their T-2/HT-2 content. We employed liquid chromatography-mass spectrometry (LC-MS) and an immunological screening method (enzyme-linked immunoabsorbant assay (ELISA)) for T-2/HT-2 determination. Detection limits were between 1 and 10 µg kg^?1 in different matrices. T-2/HT-2 concentrations determined by ELISA in oat samples from ten different agricultural sites in Germany were between 9 and 623 µg kg^?1. The median and 90th percentile were 48 and 191 µg kg^?1 T-2/HT-2, respectively. One site showed six times higher T-2/HT-2 levels than the other sites, where concentrations ranged from 322 to 623 µg kg^?1. In 80% of the samples the cultivars Pergamon and Ivory had the lowest concentration of T-2 and HT-2 toxins. Using LC-MS for T-2/HT-2 determination, cleaning of the raw material did not lead to significant reductions of T-2 and HT-2 levels, whereas de-hulling led to a reduction of over 90%. Boiling of oat meal produced from cleaned raw material to yield ‘porridge’ resulted in varying T-2/HT-2 levels in experimental replicates. No major reduction of T-2/HT-2 levels in cooked porridge was obtained. Standardized baking experiments using 20% oat meal showed that T-2 and HT-2 toxins are relatively stable during the baking process, probably due to their temperature stability.     

Survey of fumonisins B1, B2 and B3 in conventional and organic retail corn products in Spain and Italy and estimated dietary exposure

A survey was conducted to determine the occurrence of fumonisin B₁, B₂ and B₃ during 2007 in 186 samples of organic and conventional locally available corn products. Samples included baby food (n = 62), corn flour (11), cornflakes (23), pasta (14), cookies (17) and other corn products (59) were obtained from popular markets of Valencia (Spain) and Perugia (Italy). The analytical method used pressurized liquid extraction and liquid chromatography/electrospray ionization tandem mass spectrometry with a triple quadrupole (QqQ) analyser. Of the 104 Spanish samples, 22% contained levels in the range of 2–449 µg kg^?1, 2–229 µg kg^?1 and 6–105 µg kg^?1 for FB₁, FB₂ and FB₃, respectively, while 19 (23%) of the 82 Italian samples were positive with quantifiable levels between 2–235 µg kg^?1, 3–187 µg kg^?1, and 4–40 µg kg^?1 for fumonisins B₁, B₂ and B₃, respectively. Overall, none of the Italian samples and only one organic baby food sample from a Spanish market was above the maximum permitted levels established by European legislation. Fumonisins were found mostly in corn flour followed by cookies and cornflakes. Eleven samples from Spain and nine samples from Italy were organic products, being contaminated the 72% and 77% of the samples, respectively. Analysis of the results showed that levels of fumonisins in corn products were similar in Italy and Spain. The safety of fumonisin intake through corn products was demonstrated by the calculation of the estimated daily intake of both populations considering organic and conventional products separately, which ranged from 1.7 × 10^?3 to 0.72 µg kg^?1 bw day^?1 and comparing them with the provisional maximum total daily intake (PMTDI) of 2 µg kg^?1 bw day^?1 established by the European Union.     

Aflatoxin M1 contamination in cow and buffalo milk samples from the North West Frontier Province (NWFP) and Punjab provinces of Pakistan

A total of 178 milk samples (94 of buffalo and 84 of cow) were randomly taken from Punjab and the North West Frontier Province (NWFP) of Pakistan (n?=?89 in each province) and analysed for the presence of aflatoxin M1 (AFM₁) by HPLC-FLD. From Punjab about 46% of buffalo's and 49% of cow's milks were contaminated with AFM₁ as compared with 52% and 51% for milk samples from NWFP, respectively. Overall, the mean AFM₁ concentration was 0.046?µg?kg^?1 with a maximum of 0.350?µg?kg^?1. All samples complied with the Codex Alimentarius limit of 0.50?µg?kg^?1 for AFM₁ in milk, but 16.3% of samples exceeded the European Union maximum level of 0.05?µg?kg^?1. Another set of 415 buffalo's and cow's milk samples (213 morning milks and 202 evening milks) were analysed. Statistical analysis revealed significant differences (p?<?0.05) between mean AFM₁ concentrations in milk during the morning (0.043?µg?kg^?1) and the evening (0.028?µg?kg^?1) lactation times.