Development and Validation of an UHPLC-MS/MS Method for the Simultaneous Determination of 11 EU-Regulated Mycotoxins in Selected Cereals.

The availability of reliable sensitive multi-analyte methods for unambiguous determination of mycotoxins is crucial for ensuring food and feed safety, considering their adverse health effects and (co-)occurrence in various foods. Accordingly, a multi-mycotoxin confirmatory method for simultaneous determination of 11 mycotoxins regulated in cereals within the European Union (EU) using ultra-high performance liquid chromatography coupled to tandem mass spectrometry (UHPLC-MS/MS) was developed and in-house validated to fit the EU legislation requirements for analytical methods. A simple sample preparation was based on a solid−liquid extraction using a solvent mixture acetonitrile/water/formic acid (79/20/1, v/v/v) and a dilution of raw extract using water/acetonitrile/formic acid (79/20/1, v/v/v) before instrumental analysis. Average recoveries in all three validated cereal crop types (maize, wheat, and barley), spiked at multiple levels, were found acceptable for all analytes when matrix-matched calibration was used, ranging from 63.2% to 111.2% and also showing very good repeatability, with relative standard deviations below 20%. Matrix effect (SSE) evaluation revealed maize as the most complex of the three analyzed cereal matrices, with strong SSE (<50% and >150%) recorded for all 11 analyzed mycotoxins. An additional method verification was performed through successful participation in proficiency testing schemes, with the achieved z-scores generally in the acceptable range of −2 ≤ z ≤ 2. The obtained validation results demonstrated the suitability of the developed confirmatory multi-mycotoxin UHPLC-MS/MS method based on a dilute-and-shoot principle for the simultaneous determination of low concentrations of 11 EU-regulated mycotoxins in cereals, including aflatoxins B1, B2, G1 and G2, deoxynivalenol, fumonisins B1 and B2, zearalenone, T-2 and HT-2 toxins, and ochratoxin A.

Regulated Mycotoxin Occurrence and Co-Occurrence in Croatian Cereals.

A total of 209 samples of various cereal crops (maize, wheat, barley, rye and oats) grown in Croatian fields during 2016 and 2017 were collected to analyze and determine the occurrence and co-occurrence of EU regulated mycotoxins in cereals (AFB1, AFB2, AFG1, AFG2, DON, FB1, FB2, ZEA, T-2, HT-2 and OTA). The analysis, performed by a validated confirmatory LC-MS/MS method based on a dilute and shoot principle, highlighted Fusarium mycotoxins as the main contaminants, often co-occurring in samples from both years (50.0% in 2016 and 33.7% in 2017). DON was found to be the most frequent mycotoxin, present in 72.5% of the 2016 samples and 32.6% of the 2017 samples, while maize proved to be the most contaminated cereal type of both years with FUM as the most abundant mycotoxins, with an average concentration of 1180 µg/kg. Moderate temperatures with periods of high humidity favored the accumulation of DON in wheat samples instead of other Fusarium mycotoxins, while similar conditions favored maize contamination with FUM. A total of 8.3% of all the 2016 harvest samples and 7.9% of the 2017 harvest samples were assessed as non-compliant, containing mycotoxins in concentrations higher than the levels set by the EU legislation for food.

Corn silage fungal-based solid-state pretreatment for enhanced biogas production in anaerobic co-digestion with cow manure.

The objective of this research was to use white-rot fungus Trametes versicolor for corn silage pretreatment and to investigate the effect of pretreatment on biogas productivity. Semi-continuous pilot-scale experiment, comprised of two experimental phases, was carried out. In the first phase, operational conditions of the full-scale biogas plant were reproduced at pilot-scale. In that phase, the reactor was daily fed with the mixture of cow manure, digestate from industrial postfermentor, corn grits and ensiled corn silage, and the average methane generation rate was 0.167 m3CH4 kgVS-1. In the second phase, corn grits and ensiled corn silage were replaced with corn silage pretreated with T. versicolor, and the average methane generation rate increased up to 0.236 m3CH4 kgVS-1. The results of this study suggest that application of fungal-based solid-state pretreated corn silage has positive effect on pH stability and increase the biogas productivity.

Biogas Production from Brewer's Yeast Using an Anaerobic Sequencing Batch Reactor.

Renewable energy sources are becoming increasingly important in the beverage and food industries. In the brewing industry, a significant percentage of the used raw materials finishes the process as secondary resource or waste. The research on the anaerobic digestion of brewer's yeast has been scarce until recent years. One of the reasons for this is its use as a secondary resource in the food industry and as cattle feed. Additionally, market value of brewer's yeast is higher than its energy value. Due to the increase of energy prices, brewer's yeast has become of interest as energy substrate despite its difficult degradability in anaerobic conditions. The anaerobic co-digestion of brewer's yeast and anaerobically treated brewery wastewater was studied using a pilot-scale anaerobic sequencing batch reactor (ASBR) seeded with granular biomass. The experiments showed very good and stable operation with an organic loading rate of up to 8.0 kg/(m3·day), and with a maximum achieved organic loading rate of 13.6 kg/(m3·day) in a single cycle. A specific biogas productivity of over 0.430 m3/kg of the total chemical oxygen demand (COD) inserted, and total COD removal efficiencies of over 90% were achieved. This study suggests that the brewer's yeast can be successfully digested in an ASBR without adverse effects on the biogas production from brewer's yeast/wastewater mixtures of up to 8% (by volume). By using the brewer's yeast in the ASBR process, the biogas production from brewery wastewater could be increased by 50%.

Influence of Thermal and Bacterial Pretreatment of Microalgae on Biogas Production in Mesophilic and Thermophilic Conditions.

Microalgae biomass has a great potential in search for new alternative energy sources. They can be used as a substrate for the biogas production in anaerobic digestion. When using microalgae, the efficiency of this process is hampered due to the resistant cell wall. In order to accelerate the hydrolysis of cell wall and increase the efficiency of biogas production we applied two different pretreatments - biological and thermal under mesophilic and thermophilic conditions. During biological pretreatment we incubated microalgae with anaerobic hydrolytic bacteria Pseudobutyrivibrio xylanivorans Mz5T. In thermal pretreatment we incubated microalgae at 90 °C. We also tested a combined thermal and biological pretreatment in which we incubated P. xylanivorans Mz5T with thermally pretreated microalgae. Thermal pretreatment in mesophilic and thermophilic process has increased methane production by 21% and 6%, respectively. Biological pretreatment of microalgae has increased methane production by 13%, but only under thermophilic conditions (pretreatment under mesophilic conditions showed no effect on methane production). Thermal-biological pretreatment increased methane production by 12% under thermophilic conditions and by 6% under mesophilic conditions.

Anaerobic Biodegradation of Raw and Pre-treated Brewery Spent Grain Utilizing Solid State Anaerobic Digestion.

The brewery spent grain (BSG) represents approximately 85% of the total quantity of by-products from the brewing industry. The biogas production from the BSG has been the subject of several studies in recent years, due to relatively high energy consumption in the brewing process and due to the increasing energy costs. The biodegradability of raw and pre-treated BSG in a single-stage and two-stage solid-state anaerobic digestion (SS-AD) system was determined in this study. The results show that the BSG have a biogas potential of 120 L/kg(-1). In the single-stage system, the biogas yield obtained from raw BSG (87.4 L/kg(-1)) was almost equal to the yield obtained from the pre-treated BSG (89.1 L/kg(-1)), while the methane yield was 51.9 and 55.3 L/kg(-1) and the biodegradation was 62.0% and 62.2% for raw and pre-treated BSG, respectively. In two-stage SS-AD the pre-treated BSG showed better results, with the biogas yield of 103.2 L/kg(-1) and the biodegradation of 73.6%, while the biogas yield obtained from raw BSG was 89.1 L/kg(-1), with the biodegradation of 63.5%. In two-stage process the obtained methane yields from raw and pre-treated BSG were identical (58.7 L/kg(-1)).