Elisa and HPLC analyses of deoxynivalenol in maize and wheat

Deoxynivalenol (DON) is a part of the family of mycotoxins called trichothecenes which are produced by a number of different Fusarium mold species. The presence of DON in 25 wheat and 25 maize samples was examined by Enzyme Linked Immunosorbent Assay (ELISA) and High Performance Liquid Chromatography (HPLC) methods. The presence of DON was detected and determined in 5 (20%) maize and 6 (25%) wheat samples by both of the methods. Correlation between ELISA and HPLC results was established, with the correlation coefficients (r) of 0.9691 and 0.9735 for wheat and maize samples, respectively. The results obtained by ELISA method were significantly higher than those obtained by HPLC method. This fact can be explained by the presence of conjugated or masked mycotoxins in the samples, especially DON-3-glucoside (DON-3-Glc), which could not be determined by HPLC method due to the lack of external standards. Contrary to this, being insufficiently selective towards masked DON, ELISA method measures total DON content of a sample. According to the obtained results, ELISA can be used as a reliable screening method, but the confirmation of positive results must be done by HPLC method

Prisustvo zearalenona u najčešće uzgajanim sortama pšenice u Srbiji

A total of 45 samples of wheat from three different locations in Vojvodina were analyzed for the presence of zearalenone. Analytical methods based on clean-up by solid-phase extraction (SPE) columns and detection by liquid chromatography were used after validation. Limit of detection for ZEA in wheat was 18.6 μg/kg and the limit of quantification was 56.5 μg/kg. Recovery values ranged between 86% and 97%. The occurrence of ZEA in wheat was rather high with 53.3% of positive samples with the average value of 330 μg/kg. Incidences were found from 68 μg/kg to 1079 μg/kg. Contamination levels were above the established maximum limit for unprocessed cereals, other than maize, in as many as seventeen samples. These results were compared to the results of investigation of deoxynivalenol and fumonisin content, established in our previous work on the same samples. The results obtained were also compared to those of the neighboring countries where the relevant data existed and to the data of previous studies in our country.Na prisustvo zearalenona analizirano je ukupno 45 uzoraka pšenice sa tri različite lokacije u Vojvodini. Korišćene su analitičke metode zasnovane na prečišćavanju ekstrakcijom na čvrstoj fazi, te kvantifikacija tečnom hromatografijom, nakon validacije metode. Granica detekcije za zearalenon u pšenici je iznosila 18,6 μg/kg, a granica određivanja 56,5 μg/kg. Efikasnost metode je bila u opsegu od 86% do 97%. Zearalenon je bio prisutan u 53,3% ispitivanih uzoraka, sa prosečnim sadržajem od 330 μg/kg. Dobijene vrednosti sadržaja zearalenona su bile u opsegu od 68 μg/kg do 1079 μg/kg. U čak sedamnaest uzoraka je pronađena koncentracija ovog toksina koja prevazilazi maksimalni dozvoljeni sadržaj zearalenona u netretiranim žitaricama. Ovi rezultati su upoređeni sa vrednostima sadržaja deoksinivalenola i fumonizina u istim uzorcima dobijenim u našim prethodnim istraživanjima. Rezultati su takođe upoređeni sa dostupnim rezultatima dobijenim u našoj i susednim zemljama tokom prethodnih godina

Fumonisins and co-occurring mycotoxins in north Serbian corn

The presence of fumonisin has not been regulated in the legislation of the Republic of Serbia. Therefore, the data on contamination of cereals, especially corn, which is highly susceptible to contamination by this toxin, are not sufficient. This paper presents the results of testing the corn samples collected in the autumn 2009 on the territory of Bačka. Samples were analyzed for the contents of fumonisins and it was determined whether there is a correlation between the moisture content, total number and class of fungi, as well as the content of aflatoxin, ochratoxin and zearalenone. Using enzymatic immunoaffinity method it was discovered that the highest percentage of samples were contaminated with fumonisins, which was probably due to the presence of Fusarium molds as the most abundant ones. The positive samples contained fumonisin in the concentrations from 0.030 to 1.52 mg kg−1. The influence of the climate and moisture content of grain on fungal contamination and mycotoxin production was analyzed in order to investigate the predictability of the presence of mycotoxins

Prisustvo deoksinivalenola u uzorcima strnih žita u žetvenoj 2009/10. godini

Fusarium head blight (FHB) is present in all growing regions of small grains and causes significant losses in yield and grain quality. In our environmental conditions, dominant species is Fusarium graminearum Group 2. During 2009/10 there was a significant Fusarium infestation on wheat, barley and triticale. The aim of this study was to examine the contents of deoxynivalenol (DON) in cereal samples taken after 2009/10 harvest season. We analyzed 22 NS varieties of small grains from Rimski Šančevi, including 16 varieties of winter wheat, one facultative wheat variety, four varieties of winter barley and one variety of triticale. Analytical methods based on clean-up by solid-phase extraction (SPE) columns and detection by liquid chromatography were used. Fifteen out the 22 analyzed samples were positive for the presence of DON at a mean level of 0.537 mg/kg. The highest concentration was 1.952 mg/kg. These findings were in correlation with percentage of the Fusarium damaged kernels.Patogeni iz roda Fusarium su prisutni u svim rejonima gajenja strnih žita i nanose značajne gubitke u prinosu i kvalitetu zrna. Fuzariozu klasa pšenice prouzrokuje veći broj vrsta iz roda Fusarium. U našim uslovima gajenja dominantna je vrsta Fusarium graminearum Grupa 2. U toku 2009/10. godine došlo je do značajne pojave fuzarioza na pšenici, ječmu i tritikaleu. Cilj rada je bio da se ispita sadržaj DON-a na uzorcima strnih žita uzetih posle žetve iz uslova prirodne zaraze. Analizirane su 22 novosadske sorte strnih žita iz lokaliteta Rimski Šančevi, od čega: 16 sorti ozime pšenice, jedna fakultativna sorta pšenice, četiri sorte ozimog ječma i jedna sorta tritikalea. Uzorci su uzeti sa parcela različitih površina u zavisnosti od značaja i rasprostranjenosti gajene sorte. Analitičko određivanje je zasnovano na prečišćavanju sirovog ekstrakta analiziranih uzoraka pomoću tzv. Mucosep kolona, a zatim je sadržaj DON-a kvantitativno određen tečnom hromatografijom. Od 22 analizirana uzorka strnih žita čak 15 (68,2%) je bilo pozitivno na prisustvo DON-a. Još veći procenat zaraženosti DON-om je utvrđen kada je u pitanju samo pšenica (82,4%). Prosečan sadržaj DON-a je iznosio 0,537 mg/kg a najveća koncentracija je utvrđena u uzorku tritikalea i iznosila je visokih 1,952 mg/kg. Od svih uzoraka koji su bili pozitivni na prisustvo ovog mikotoksina, 2 su prevazilazila koncentracije koje su propisane od strane Evropske komisije. Sve ovo ukazuje na visoku zaraženost strnih žita sa naših polja iz žetve 2010. Procenat fuzarioznih zrna kod 16 ispitivanih sorti pšenice kretao se od 1 do 11,5%, a gubici u masi 1000 zrna od 1,2 do 5,7%. Između jačine zaraze u polju i sadržaja mikotoksina DON ustanovljena je potpuna pozitivna korelacija kod pojedinih sorti. Fakultativna sorta pšenice Nataša je imala visoku koncentraciju DON od 1,572 mg/kg, pri stepenu zaraze u polju od 33,3% zaraženih klasova po 1 m2. Sorta Zvezdana imala je najniži procenat zaraze u polju od 1% i kod nje nije determinisano prisustvo mikotoksina DON

Potencijal za proizvodnju fumonizina kod izolata Fusarium verticillioides sa zrna kukuruza i pšenice iz Srbije

The production of fumonisins by potentially toxigenic Fusarium verticillioides isolates originating from Serbian maize and wheat kernels was tested in vitro. A total of six F. verticillioides isolates were incubated on yeast extract sucrose medium (YESA) for 4 weeks at 25 °C in the dark. Their toxin production potential was tested by applying a modified HPLC method for determination of fumonisins in cereals, since the TLC method gave no results. Analyses were performed on a HPLC-FLD system after sample extraction from YESA and extract clean-up on a SPE column. Although the isolates were tested for fumonisin B1, B2 and B3, only fumonisin B1 was detected. The results showed that all tested isolates had toxigenic potential for fumonisin B1 production. The average fumonisin B1 production of the isolates ranged from 7 to 289 µg/kg, thus indicating a highly variable toxigenic potential among the isolates. Isolate 1282 expressed the highest toxigenic potential for fumonisin B1 production (289 µg/kg), while isolate 2533/A showed a questionable potential for fumonisin production (7 µg/kg).Proizvodnja fumonizina kod potencijalno toksikogenih Fusarium verticillioides izolata sa zrna kukuruza i pšenice poreklom iz Srbije testiran je in vitro. Ukupno šest izolata F. verticillioides inkubirano je na agarizovanoj podlozi sa ekstraktom kvasca i saharozom (YESA) tokom četiri nedelje, u mraku na 25 °C. Kapaciteti odabranih izolata za proizvodnju toksina detektovani su primenom modifikovane HPLC metode za određivanje fumonizina u zrnu žitarica, jer brza trijažna TLC metoda nije dala pozitivne rezultate. Analiza je izvedena na HPLC-FLD sistemu nakon ekstrakcije uzorka iz YESA kultura izolata i prečišćavanja ekstrakta na SPE koloni. Sve analize su urađene u tri ponavljanja. Iako su izolati F. verticillioides testirani na prisustvo fumonizina B1, B2 i B3, samo je fumonizin B1 bio detektovan. Rezultati su pokazali da svi testirani izolati imaju potencijal u sintezi fumonizina B1. Prosečna vrednost proizvedenog fumonizina B1 kod izolata F. verticillioides kretala se od 7 do 289 µg/kg, što ukazuje na izuzetno varijabilan toksigeni potencijal istih izolata. Izolat označen sa 1282 pokazao je najveći potencijal za biosintezu fumonizina B1 (289 µg/kg), dok je izolat 2533/A ispoljio diskutabilan potencijal za proizvodnju istog fumonizina (7 µg/kg)

Vascular and upper gastrointestinal effects of non-steroidal anti-inflammatory drugs: meta-analyses of individual participant data from randomised trials

Summary Background The vascular and gastrointestinal eff ects of non-steroidal anti-infl ammatory drugs (NSAIDs), including selective COX-2 inhibitors (coxibs) and traditional non-steroidal anti-infl ammatory drugs (tNSAIDs), are not well characterised, particularly in patients at increased risk of vascular disease. We aimed to provide such information through meta-analyses of randomised trials. Methods We undertook meta-analyses of 280 trials of NSAIDs versus placebo (124 513 participants, 68 342 personyears) and 474 trials of one NSAID versus another NSAID (229 296 participants, 165 456 person-years). The main outcomes were major vascular events (non-fatal myocardial in farction, non-fatal stroke, or vascular death); major coronary events (non-fatal myocardial infarction or coronary death); stroke; mortality; heart failure; and upper gastrointestinal complications (perforation, ob struction, or bleed). Findings Major vascular events were increased by about a third by a coxib (rate ratio [RR] 1·37, 95% CI 1·14-1·66; p=0·0009) or diclofenac (1·41, 1·12-1·78; p=0·0036), chiefl y due to an increase in major coronary events (coxibs 1·76, 1·31-2·37; p=0·0001; diclofenac 1·70, 1·19-2·41; p=0·0032). Ibuprofen also signifi cantly increased major coronary events (2·22, 1·10-4·48; p=0·0253), but not major vascular events (1·44, 0·89-2·33). Compared with placebo, of 1000 patients allocated to a coxib or diclofenac for a year, three more had major vascular events, one of which was fatal. Naproxen did not signifi cantly increase major vascular events (0·93, 0·69-1·27). Vascular death was increased signifi cantly by coxibs (1·58, 99% CI 1·00-2·49; p=0·0103) and diclofenac (1·65, 0·95-2·85, p=0·0187), nonsignifi cantly by ibuprofen (1·90, 0·56-6·41; p=0·17), but not by naproxen (1·08, 0·48-2·47, p=0·80). The proportional eff ects on major vascular events were independent of baseline characteristics, including vascular risk. Heart failure risk was roughly doubled by all NSAIDs. All NSAID regimens increased upper gastrointestinal complications (coxibs 1·81, 1·17-2·81, p=0·0070; diclofenac 1·89, 1·16-3·09, p=0·0106; ibuprofen 3·97, 2·22-7·10, p<0·0001; and naproxen 4·22, 2·71-6·56, p<0·0001). Interpretation The vascular risks of high-dose diclofenac, and possibly ibuprofen, are comparable to coxibs, whereas high-dose naproxen is associated with less vascular risk than other NSAIDs. Although NSAIDs increase vascular and gastrointestinal risks, the size of these risks can be predicted, which could help guide clinical decision making. Funding UK Medical Research Council and British Heart Foundation. Introduction Non-steroidal anti-infl ammatory drugs (NSAIDs) are among the most widely used drugs in the world. They are chiefl y used to treat pain, but their long-term use is limited by serious gastrointestinal side-eff ects. NSAIDs inhibit the two recognised forms of prostaglandin G/H synthase (also referred to as cyclo-oxygenase [COX]), namely COX-1 and COX-2. 1 Since the analgesic and antiinfl ammatory eff ects of NSAIDs are mediated by inhibition of COX-2, and their gastrointestinal side eff ects mostly by inhibition of COX-1, NSAIDs which selectively inhibit COX-2 might reduce the risk of gastrointestinal toxicity compared with other NSAIDs. Several such COX-2 selective drugs (collectively known as coxibs) were developed in the 1990s, and early trials comparing coxibs versus traditional NSAIDs (tNSAIDS) seemed to confi rm that coxibs at doses with similar analgesic effi cacy had less gastrointestinal toxicity. 2,3 Unfortunately, however, subsequent placebo-controlled trials also showed unequivocally that coxibs were associated with an increased risk of atherothrombotic vascular events. 4,5 Soon after these placebo-controlled trials were reported, a meta-analysis of randomised trials comparing a coxib versus placebo or a coxib versus tNSAID indicated that some tNSAIDs might also have adverse eff ects on atherothrombotic events, but that these hazards might depend on the degree and duration of suppression of platelet COX-1

Which clinical variables have the most significant correlation with quality of life evaluated by SF-36 survey in Croatian cohort of patient with ankylosing spondylitis and psoriatic arthritis?

The aim of our study was to assess clinical variables with the best correlation to quality of life (QOL) assessed by medical outcome survey Short-Form 36 (SF-36) in patients with spondyloarthritides, including ankylosing spondylitis (AS) and psoriatic arthritis (PsA). We analyzed the cohort of 54 patients (22 patients with PsA and 32 patients with AS), who filled the Croatian version of SF-36. For each type of arthritis, patients were clinically evaluated using the extensive list of clinical variables categorized into subjective and objective group. For AS patients, subjective and objective variables (spinal mobility measurements, clinical assessment of spinal pain, patient assessments of disease activity and pain) correlated mainly with the physical functioning concept of SF-36. Patients assessments of fatigue correlated with the energy/fatigue subscale, whereas patient assessment of enthesial pain correlated with the pain subscale. Correlations between clinical variables and SF-36 concepts of PsA patients showed more diverse distribution than for AS. Objective variables (spinal mobility measurements, a 76-joint score, clinical assessment of spinal pain) correlated with concepts concerning physical health and pain. Several subjective patient assessments correlated with energy/fatigue, emotional well-being, pain and general health subscales. Both patient and physician assessment of PsA activity correlated with the role limitations due to emotional problems. Bath ankylosing spondylitis functional index (BASFI) had the strongest correlation with the physical functioning concept of SF-36 in both diseases. Our findings provide important information to help selecting the variables with strongest impact on QOL, for better planning the management strategies and achieving better rehabilitation results

Validation and application of FTIR spectroscopy in raw milk analysis

The aim of this study was to investigate whether FTIR spectroscopy is an accurate and valid technique for the assessment of quality parameters in raw cow's milk: fat, protein, lactose, and total solids. The assessment was based on calibration series and comparison with reference material. Furthermore, it takes into account the results obtained in the inter-laboratory comparisons (proficiency testing). The calibration samples were purchased from the accredited regional reference laboratories. The validation parameters included linearity, accuracy, repeatability, reproducibility, and robustness. The linearity ratio was 0.95%. The biases calculated for the fat, protein, lactose and dry matter were -0.33, 0.31, -0.25, and 0.06 respectively. The F value from the F-test was used to determine the significant differences between two independent sets of the results. The obtained results were as follows: 1.469 for fat, 1.634 for protein, 1.192 for lactose, and 0.528 for dry matter. The intra-laboratory reproducibility calculated as the Horwitz Ratios for all parameters were within the criterion limits (0.5 to 0.8). The data obtained for carry-over were 0.27% for fat, 0.52% for protein, 0.47% for lactose, and 0.47% for dry matter. Based on the obtained results it can be concluded that the FTIR spectroscopy is a reliable instrumental technique for the determination of fat, protein, lactose and total solids in raw cow's milk

Deoxynivalenol occurrence in Serbian maize under different weather conditions

The aim of this paper was to investigate deoxynivalenol (DON) occurrence in maize samples originating from two harvest seasons in Serbia. The key differences between harvest seasons were weather conditions, specifically the humidity. The samples were analyzed using high performance liquid chromatography with DAD detection, after clean-up on SPE columns. In samples from 2014, DON was found in 82 (100.0%) samples with the average content of 2.517 mg/kg (ranged from 0.368 to 11.343 mg/kg). Two samples exceeded maximum level permitted by EU regulations. However, analyzing larger number of samples (163) from 2015 harvest season, DON was present in 51 (31.3%) samples in significantly lower concentrations (average of 0.662 mg/kg, ranged from 0.106 to 2.628 mg/kg). None of the samples from 2015 exceeded maximum level permitted by EU regulations. The data on DON presence in Serbian maize were in relation to the different weather conditions that prevailed during the two harvest seasons. [Project of the Serbian Ministry of Education, Science and Technological Development, Grant no. 172042