Determination of ochratoxin A in domestic and imported beers in italy by immunoaffinity clean-up and liquid chromatography

A method first developed to quantify ochratoxin A in wine has been applied to the analysis of domestic and imported beers in Italy. The method uses commercial immunoaffinity columns for clean-up and high-performance liquid chromatography for quantification of the toxin. Beer was degassed, then diluted with a polyethylene glycol-sodium hydrogencarbonate solution and applied to an OchraTest immunoaffinity column. Ochratoxin A was eluted from the immunoaffinity column with methanol and quantified by reversed-phase HPLC with fluorometric detector. Average recoveries of ochratoxin A from blank beer spiked at levels from 0.04 to 1.0 ng/ml ranged from 93.8% to 100.4%, with relative standard deviations between 3.3% and 5.7%. The detection limit was 0.01 ng/ml based on a signal-to-noise ratio of 3:1. The analysis of 61 samples of domestic (10) and imported (51) beers showed ochratoxin A levels ranging from <0.01 to 0.135 ng/ml with an incidence of contamination of 50% and no substantial difference between strong and pale beers.     

Detection ofFusarium trichothecenes (nivalenol,deoxynivalenol, fusarenone and 3-acetyldeoxynivalenol) by high-performance liquid chromatography

Summary Separation and determination ofFusarium trichothecenes: nivalenol, deoxynivalenol, fusarenone and 3-acetyldeoxynivalenol, were carried out by highperformance liquid chromatography with UV detection, and methanol—water (30/70) as mobile-phase on Lichrosorb RP-18 column. Detection limits were 2, 3, 3 and 5ng per injection of nivalenol, deoxynivalenol, fusarenone and 3-acetyldeoxynivalenol, respectively. The method appears to be adequate for detection of the above trichothecenes in contaminated corn and rice as well as in cultures ofFusarium spp. A preliminary TLC purification was necessary for detection of fusarenone in corn.     

Separation of chlamydosporol epimers by reversed-phase HPLC using commercial solvent optimisation software

Summary Two epimers of the mycotoxin chlamydosporol were separated by HPLC on an RP-18 column using a quaternary mobile phase consisting of water (79.1%), methanol (10.0%), acetonitrile (10.4%) and tetrahydrofuran (0.5%), with a flow rate of 1 ml min^–1. This optimal composition of mobile phase, with which the resolution value for the two epimers (1 and2) was 2.73 with retention times of 5.88 and 7.12 min, respectively, was achieved by the application of Philips Solvent Optimisation Software PU 6100. The presence of free silanols on the stationary phase was shown to be an essential requirement for the separation of the chlamydosporol epimers.     

Discrimination between ethanol inhibition models in a continuous alcoholic fermentation process using flocculating yeast

Discrimination between different rival models for describing the inhibitory effect of ethanol both on yeast growth and on fermentation was studied for a continuous process of alcoholic fermentation in a tower reactor with recycling of flocculating cells. Models tested include linear, parabolic, hyperbolic, exponential, and generalized nonlinear power-law types. The best expressions were identified under the criteria that all the kinetic parameters should assume acceptable values in a feasible range and should result in the best fit of the experimental data. The kinetic parameters were estimated from steady-state data of several sugar concentrations in feeding stream (S₀ = 160, 170, 180, 190, 200 g/L), constant dilution rate (D = 0.2 h^-1), recycle ratio (α = 13.6), and temperature (T = 30°C). The best model for the yeast growth was of power-law type, whereas for the product formation the best model was of linear type. These models were able to reproduce the trends of the process variables satisfactorily.     

Selective determination of altertoxins by high-performance liquid chromatography with electrochemical detection with dual "in-series" electrodes

A selective method for the determination of altertoxin-I and altertoxin-II by high-performance liquid chromatography with electrochemical detection is described. Altertoxins were separated on a reversed-phase column with methanol-water containing 0.1 M sodium nitrate and 1 mM nitric acid (60:40) as eluent and detected with dual in-series electrodes operating in the "redox" mode (generator electrode +1.0 V, indicator electrode -0.1 V). The method was applied successfully to the determination of sub-ppm levels of altertoxins in samples of maize, rice and tomatoes infected by Alternaria alternata.     

Mycotoxin profile of Fusarium langsethiae isolated from wheat in Italy: production of type‐A trichothecenes and relevant glucosyl derivatives

Fusarium langsethiae, formally described as a new species over a decade ago, has been identified as the main producer of HT‐2 (HT2) and T‐2 (T2) toxins in Europe in small cereal grains. Mycotoxin contamination caused by this Fusarium species can represent a food safety hazard that deserves further attention. In the present work, the mycotoxin profile in wheat cultures of F. langsethiae is presented with particular reference to the production of major type‐A trichothecenes and their glucosyl derivatives. F. langsethiae isolates, representative of the major Italian wheat cultivation areas, were tested for the production of T2, HT2, diacetoxyscirpenol (DAS) and neosolaniol (NEO), and relevant glucosyl derivatives. Liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) was used for the identification and chemical characterization of these metabolites. F. langsethiae isolates under investigation resulted to be potent producers of T2, HT2 and NEO. Furthermore, a well‐defined set of isolates, all originating from Central Italy, produced also DAS. All isolates were found to be able to produce HT2 glucosyl derivatives, whereas only traces of T2 glucoside were detected in one sample. Furthermore, two mono‐glucosyl derivatives of NEO and one mono‐glucoside derivative of DAS were identified and characterized. The screening for the presence/absence of glucosylated trichothecenes in analyzed fungal extracts revealed a general co‐occurrence of these derivatives with the parent toxin at levels that could be roughly estimated to account up to 37% of the relevant unconjugated toxin. This is the first report of the production of glucosylated trichothecenes by F. langsethiae cultured on small grains. Copyright © 2013 John Wiley & Sons, Ltd.     

Experimental design for in-house validation of a screening immunoassay kit. The case of a multiplex dipstick for Fusarium mycotoxins in cereals

The benefits of using rapid qualitative methods to verify compliance of food and feed with legislation requirements include user-friendly format, the possibility of detection without expensive instrumentation, rapid response and affordable price. Prior to their use, however, the methods have to pass validation experiments, in order to assess their performance profile. An experimental protocol for in-house validation of a screening immunoassay has been designed and applied to evaluate performance characteristics of a multiplex dipstick kit for the determination of major Fusarium toxins, namely zearalenone, T-2 and HT-2 toxins, deoxynivalenol and fumonisins in wheat and maize. The test is intended for screening of cereals on the presence/absence of these mycotoxins at maximum permitted levels established by European legislation or target levels. The response of the measurement is determined with a reader device. Samples classified as negative are considered as compliant, whereas positive samples need to be re-analysed with confirmatory methods. The in-house validation design consisted of three steps, namely (1) estimating the precision of the method including "between day" effects and influences from different varieties of the matrices, (2) establishing robust cutoff values for the dipstick response at target mycotoxin levels assuming an acceptable rate of false negative results of 5 % and (3) assessment of the rate of false positive results of blank samples and samples containing the target analytes below the legal limits. The total precision expressed as relative standard deviation and determined individually for each analyte/concentration/matrix combination varied from 9 to 30 % and was considered as acceptable. In 17 out of 28 cases, the repeatability standard deviation was the most important factor. The predominance of the repeatability compared to the other factors (matrix, days) was an indicator for the ruggedness of the assay. The validation study demonstrated that the test was able to differentiate blank samples from samples contaminated at target mycotoxin levels with a false positive rate lower than 6 %. Considering realistic mycotoxin occurrence in European samples, significant economical benefits can be expected when using the test under real-world conditions.

A two-dimensional model of the distribution of trace gases in the stratosphere and troposphere

Summary We report the results of computations of the distribution of trace gases in the troposphere and stratosphere with a two-dimensional photochemical transport model. The region covered by the model extends from pole to pole and from 0 to 50 km. Atmospheric transport is included by using seasonal averaged values for mean meridional and vertical winds and eddy diffusion. The compounds transported in the model are N₂O, NO _x (NO+NO₂), NO _y (NO_x+HNO₃), N₂O₅, H₂O, CH₄ and H₂O₂. Odd hydrogen (OH+HO₂+H) is assumed in photoequilibrium. A simple scheme is proposed to simulate the condensation of H₂O and scavenging of other gases. Rayleigh scattering is included in the calculation of the photodissociation rates. The resulting distributions are compared whenever possible with experimental data. We attempt to discuss the stratospheric budget of water vapour taking into account photochemical sources (e.g. CH₄ oxidation) and sinks. It is shown that similarly to other models serious disagreement seems to exist between the calculated and observed mixing ratios of NO _x , NO _y and the ratios of the different compound within the family. The amount of experimental data is, however, too sparse to draw any definite conclusion.

---

Riassunto Si riportano i rusultati sulla distribuzione di gas minori nella stratosfera e troposfera, ottenuti con un modello bidimensionale, fotochimico con trasporto. Il modello si estende da 0 a 50 km in quota e da polo a polo in latitudine. Del trasporto si tiene conto usando i valori medi stagionali dei venti meridionali e verticali e dei coefficienti di diffusione turbolenta. Il modello tratta il trasporto per i seguenti composti: N₂O, NO _x (NO+NO₂), NO _y (NO _x +HNO₃), N₂O₅, H₂O, CH₄ e H₂O₂. L'idrogeno dispari (H+OH+HO₄) è calcolato all'equilibrio fotochimico. Un semplice meccanismo è proposto per simulare la condensazione del vapor d'acqua e di altri gas. I coefficienti di fotodissociazione sono calcolati tenendo conto dello scattering di Rayleigh. Le distribuzioni ottenute per i vari gas sono confrontate, ove ciò è possible, con i dati sperimentali. Si discute inoltre il bilancio stratosferico del vapor d'acqua tenendo conto delle sorgenti fotochimiche (cioè l'ossidazione del CH₄) e dei pozzi. Si conclude che, analogamente ad altri modelli, esistono delle importanti differenze fra i rappporti di mescolamento calcolati e qualli osservati di NO _x e NO _y . I dati sperimentali sono comunque ancora troppo scarsi per arrivare a conclusioni definitive.

---

Резюме Мы приводим результаты вычислений распределения прослеживаемых газов в тропосфере и стратосфере с помощью двумерной фотохимической модели переноса. Область, которую покрывает модель, распостраняется от полюса до полюса и от нуля до 50 км. Атмосферный перенос включает сезонные усредненные значения для средних меридиональных и вертикальных ветров и диффзию вихрей. Переносимые соединения в этой модели являются: N₂O, NO _x (NO+NO₂), NO _y (NO _x +HNO₃), N₂O₅, H₂O, CH₄ и H₂O₂. Предполагается, что нечетный водород (OH+HO₂+H) находится в фоторавновесии. Предлагается простая схема для моделирования конденсации H₂O и удаления другин газов. При вьчислении интенсивностей фотодиссоциации учитывается релеевское рассеяние. Полученные распределения сравниваются с имеющимися экспериментальными данными. Мы пытаемся обсудить стратосферный бюджет водяных паров, учитывая фотохимические источники (т.е. окисление CH₄) и стоки. Показывается, что аналогично другим моделям, по-видимому, существует существенное расхождение между вычисленными и измеренными отношениями перемешивания NO _x , NO _y и отношениями для различных соединений. Однако экспериментальные данные являются слишком разбросанными, чтобы сделать определенные выводы.