Determination of zearalenone and its metabolites α- and β-zearalenol in beer samples by high-performance liquid chromatography–tandem mass spectrometry

A fast, robust and sensitive LC–MS–MS method for the determination of zearalenone (ZON) and its metabolites α-zearalenol (α-ZOL) and β-zearalenol (β-ZOL) in beer samples is described. Sample preparation was performed by direct RP-18 solid-phase extraction of undiluted beer samples followed by selective determination of analytes by LC–MS–MS applying an atmospheric pressure chemical ionization (APCI) interface. Using the negative ion mode limits of determination of 0.03–0.06 μg l⁻¹ beer and limits of quantification of 0.07–0.15 μg l⁻¹ beer were achieved, which was distinctly more sensitive than in the positive ion mode. Twenty-three beer samples from different countries, produced from different grains and under different brewing conditions, were investigated by this method, but only in one sample could β-ZOL and ZON be detected. Independently of the type of beer, relative standard deviations between 2.1% and 3.3%, a linear working range of 0.15 μg l⁻¹ to 500 μg l⁻¹ beer and recovery rates around 100% could be achieved when zearalanone (ZAN) was used as internal standard.     

Influence of absolute pressure on mass transfer kinetics in beer dialysis

Summary Using experimental results of chemical composition of initial beer, dialysed beer and dialysate, obtained in laboratory scale equipment for dialysis, basic parameters for mass transfer kinetics were determined. It was found that absolute pressure in the system under the medium flow rate of 300 ml/min increased alcohol separate selectivity. As the result of the specific two-phase character of beer, the working pressure in the system for beer dialysis did not fall below 3–4 bar.     

Size selectivity of aqueous pores in stomatous cuticles of <Emphasis Type="Italic">Vicia faba</Emphasis> leaves

Size selectivity of aqueous pores in Vicia leaf cuticles was investigated by measuring the penetration of calcium salts into the abaxial surface of detached leaves. Molecular weights of salts ranged from 111 g mol^–1 to 755 g mol^–1. Penetration in light at 20°C and 100% humidity was a first order process and rate constants of penetration ranged from 0.39 h^–1 (CaCl₂) to 0.058 h^–1 (Ca-lactobionate). Penetration was a first order process in the dark as well, but the rate constants were smaller by a factor of 1.82. Plotting logarithmatised rate constants versus anhydrous molecular weights resulted in straight lines both in light and in the dark. The slopes per hour were very similar and the average slope was –1.2×10^–3 mol g^–1. Hence, size selectivity was not affected by stomatal opening, and in light or darkness permeability of Vicia cuticles decreased by a factor of 2.9 when molecular weight increased from 100 g mol^–1 to 500 g mol^–1. Silver nitrate was preferentially precipitated as silver chloride in guard cells, glandular trichomes and at the base of trichomes. It was concluded that these precipitates mark the location of aqueous pores in Vicia leaf cuticles. The size selectivity of aqueous pores in Vicia leaf cuticles is small compared to that observed in poplar leaf cuticles, in which permeability decreased by a factor of 7–13 for the same range of molecular weights. It is also much smaller than size selectivity of the lipophilic pathway in cuticles. These findings suggest that active ingredients of pesticides, growth regulators and chemical inducers with high molecular weights penetrate leaves at higher rates when formulated as ions.     

Grape skins as a natural support for yeast immobilization

Grape skins were used to immobilize Saccharomyces cerevisiae. In repeated batch fermentations of grape by immobilized and free cells, the maximum specific rate of alcohol production on glucose decreased from 7.98 h^–1 at 25 °C to 0.7 h^–1 at 5 °C. The rate was approximately twice as high as that on fructose. The rates for free cells were very low. The maximum alcohol yield (0.45 g g^–1) was obtained at 5 °C when the immobilized biocatalyst was used.     

The influence of flow rate and the composition of supplied CO2/air mixtures on discontinuous growth of Tetraselmis sp.

Summary This paper studies the influence of the flow rate of gaseous mixtures on the kinetics of growth and the fatty acid composition of Tetraselmis sp. at CO₂/air ratios of 3 × 10^–4 and 2 × 10^–5. The specific growth rate rises with increased flow rate up to values of approximately 0.086 h^–1 and 0.063 h^–1 at CO₂/air ratios of 3 × 10^–4 and 2 × 10^–5 respectively, when the flow rate is approximately 3 v/v per minute. At higher flow rates, the specific growth rate decreases. The polyunsaturated fatty acid content decreases slightly as the gaseous mixture flow rate increases, whereby the ratio 3/6 remains between 2 and 3, indicating good nutritional values. Offprint requests to: E. Molina     

Immobilization of invertase on a new type of macroporous glycidyl methacrylate

Invertase was immobilized via its carbohydrate moiety. The immobilized enzyme has a specific activity of 5500 IU g^–1, with 45% activity yield on immobilization. In a packed bed reactor, 90% 2.5 M sucrose was converted at a flow rate of 4 bed volumes h^–1. The obtained specific productivity at 40 °C of 3 kg l^–1 h^–1 is the best one so far. Long-term stability was 290 days in 2.5 M sucrose at 40 °C and at a flow rate of 3 bed volumes h^–1.     

Biooxidation of ferrous iron by immobilized Acidithiobacillus ferrooxidans in poly(vinyl alcohol) cryogel carriers

PVA-cryogels entrapping about 10⁹ cells of Acidithiobacillus ferrooxidans per ml of gel were prepared by freezing-thawing procedure, and the biooxidation of Fe²⁺ by immobilized cells was investigated in a 0.365 l packed-bed bioreactor. Fe²⁺ oxidation fits a plug-flow reaction model well. A maximum oxidation rate of 3.1 g Fe²⁺ l^–1 h^–1 was achieved at the dilution rate of 0.4 h^–1 or higher, while no obvious precipitate was determined at this time. In addition, cell-immobilized PVA-cryogels packed in bioreactor maintained their oxidative ability for more than two months under non-sterile conditions. Nomenclature: C _A0 – Concentration of Fe²⁺ in feed stream (g l^–1) C _A – Concentration of Fe^{2 +} in outlet stream (g l^{– 1}) D – Dilution rate of the packed-bed bioreactor (h^–1) F – Volumetric flow rate of iron solution (l h^–1) F _A0 – Mass flow rate of Fe²⁺ in the feed stream (g h^–1) K – Kinetic constant (l l^–1 h^–1) r _A – Oxidation rate of Fe²⁺ (g l^–1 h^–1) V – Volume of packed-bed bioreactor (l) X _A – Conversion ratio of Fe²⁺ (%)     

Effect of controlled substrate feeding on butyric acid production byClostridium tyrobutyricum

Summary Production of butyric acid from wheat flour hydrolysate was studied withClostridium tyrobutyricum. The mode of substrate supply was found a key parameter for fermentation performance as large improvements were obtained by feeding with a non-limiting supply of substrate. With this procedure, increases in product concentration and productivity but also in selectivity and yield for butyrate were obtained. Substrate feeding controlled by the rate of gas production was found preferable to constant rate feeding for reason of convenience and flexibility. In these conditions, a butyrate concentration of 62.8 gl^–1 was obtained with a productivity of 1.25 gl^–1 h^–1, a selectivity of 91.5% and a yield of 0.45 g per g of glucose.     

Continuous beer fermentation by high cell-density culture of bottom brewer's yeast

Continuous beer production was investigated in a high cell-density culture system which consisted of two stages for the fermentation and sedimentation of yeast cells. The continuous culture was carried out for a fermentation time of 5,500 h without contamination, at varying dilution rates and fermentation temperatures in the ranges of 0.017-0.033 h⁻¹ and 6.5–8.5°C, respectively. This process was found to be suitable for continuous and stable beer brewing. Under these conditions, the cell concentration in the first stage was about 80 times as high as that in the exit of the second stage. Concentrations of viable cells, sugar and ethanol were maintained at 1.3 × 10⁹ cells/ml, 25 and 36 g/l, respectively, and were hardly affected by fermentation temperature. Concentrations of ethyl acetate, isoamyl alcohol and isoamyl acetate were similar in the fermentation temperature ranges of 6.5–8.5°C, and the amounts at a fermentation temperature of 7°C were comparable to those of lager-type beer. Diacetyl flavor, which is known to be an effluent component that causes deterioration in the second stag e (young beer), was maintained at 1.2 ppm at a dilution rate and fermentation temperature of 0.022 h⁻¹ and 7°C, respectively. The diacetyl flavor was due to the accumulation of vicinal diketone, the precursor of which is acetohydroxy acid. The acetohydroxy acid was converted to vicinal diketone by pretreatment at 60°C for 30 min. The vicinal diketone was then consumed by the yeast during after-fermentation at a fermentation temperature of 3°C. Using this method, total vicinal diketone decreased below 0.3 ppm for an after-fermentation time of 6.8 h, which was 225 times as fast as that of after-fermentation without the pretreatment. This process may make it possible to achieve continuous beer fermentation from the fermentation stage to after-fermentation for diacetyl removal.     

Adaptation of renal function to hypotonic medium in the winter flounder (Pseudopleuronectes americanus)

Summary The kidneys of winter flounders transferred to hypotonic medium were investigated for glomerular and tubular handling of fluid and electrolytes and for the urinary excretion of proteins. Media were sea water (925 mosm·kg^–1) and brackish water (70 mosm·kg^–1).In sea water, the urine was hypertonic to the plasma in 7 fish of this study. Urine flow rate was correlated with the GFR. After adaptation to brackish water a delay of 1 to 3 days was observed until the kidneys switched from fluid retention to the excretion of large amounts of dilute urine. GFR and urine flow rate were increased from 0.61±0.08 to 1.58±0.29 ml·h^–1·kg^–1 and from 0.14±0.02 to 0.68±0.08 ml·h^–1·kg^–1, respectively . With increased filtered load the tubular reabsorption of fluid decreased from 74±2.4% to 45±11.2%. The excretion rates of sodium and potassium were increased due to decreased fractional sodium and potassium reabsorption. The urinary excretion of divalent cations, however, was reduced because the net tubular reabsorption of calcium was increased and the net secretion of magnesium was diminished.Both the urinary total protein concentration and the protein pattern showed no significant change, but the rate of protein excretion was increased from 0.21±0.04 to 0.60±0.05 mg·h^–1·kg^–1. The comparison of protein patterns obtained from urine and serum samples revealed that high molecular weight (HMW) proteins prevail in the serum whereas low molecular weight (LMW) proteins dominate in the urine. The diminished quantity of the HMW-protein fraction in the urine thus may reflect size selectivity of the glomerular filtration barrier for serum proteins also in the winter flounder.Abbreviations BW brackish water - SW sea water - GFR glomerular filtration rate - HMW heigh molecular weight - LMW low molecular weight     

Control of recombinant human endostatin production in fed-batch cultures of Pichia pastoris using the methanol feeding rate

Endostatin is a 20 kDa carboxyl-terminal fragment of collagen XVIII that strongly inhibits angiogenesis and tumor growth. The methylotrophic yeast, Pichia pastoris, is a robust expression system that can be used to study methods to improve the yields of rhEndostatin. We expressed rhEndostatin in P. pastoris under the control of the alcohol oxidase 1 (aox 1) promoter (Mut⁺ phenotype) as a model, and used a cell biomass of about 50 g l^–1 dry cell wt as a starting point for the induction phase and varied the methanol feed rate at 8 ml l^–1 h^–1, 11 ml l^–1 h^–1 and 15 ml l^–1 h^–1. While the cell growth rate was proportional to the rate of methanol delivery, protein production rate was not. These findings could be used to guide parameters for large-scale production of recombinant proteins in the P. pastoris system.     

Influence of operational parameters in the flocculation and production ofLactobacillus plantarum

Summary The influence of different operational parameters, such as the dilution rate (D) and the bleeding rate (B), in the production of a flocculent strain ofLactobacillus plantarum was studied. The effect of the dilution rate was demonstrated to be related to the lactic acid concentration inside the reactor. The effect of the bleeding rate was shown to be critical in the stabilization of the operation (due to a better pH control). It also allowed a continuous recovery of cells outside the reactor. Viability testing of the lactic starter cultures showed that operation with cell purge increased the viability of the starter cultures obtained.Nomenclature B Bleeding rate, h^–1 - D Dilution rate, h^–1 - F Feed flow rate, L h^–1 - I Feed velocity, m h^–1 - Specific growth rate, h^–1 - v Lactic acid specific productivity, g g^–1 h^–1 - P Product concentration (lactic acid), g L^–1 - P _out Product concentration leaving the system, g L^–1 - Q _b Bleeding flow rate, L h^–1 - R Recirculation velocity, m h^–1 - S Substract concentration, g L^–1 - t Time, h - T _p Time of ascensional flow (length of the column/total ascensional velocity), h - T _r Residence time (1/D), h - V Volume of the reactor, L - X Cell concentration, g L^–1 - X _out Cell concentration leaving the system, g L^–1     

Immobilization of yeast cells in PVA particles for beer fermentation

In this study, the potential of application of non-aggressive LentiKat^® technique for brewer's yeast immobilization on polyvinyl alcohol was assessed. High cell loads of about 10⁹ cells/ml were achieved by this procedure and immobilization procedure had no adverse effect on cell viability. The stability and activity of obtained immobilized biocatalyst was tested in the growth studies and fermentations. Immobilized cells exhibited high fermentation activity in both, laboratory and pilot-scale fermentations. In three successive gas-lift reactor fermentations the apparent attenuation of around 80% was reached after only 2 days, indicating good potential of immobilized cells for development of continuous primary beer fermentation. LentiKat^® particles showed high mechanical and fermentative stability, since they endured 30 days of operating time during 6-month period without significant change of cell activity, particle shape and particle size.     

Influence of oxygen limitation on glucose metabolism in Hanseniaspora uvarum K5 grown in chemostat

Summary Growth and metabolite formation were studied as a function of oxygen feed rate, in glucose-limited chemostat cultures of Hanseniaspora uvarum K₅ at a dilution rate of 0.26 h^–1. Alcoholic fermentation occured at an oxygen feed rate of 80 mmol.l^–1.h^–1 . Below this value, pyruvate decarboxylase and alcohol dehydrogenase were present at high levels. In contrast, activities of oxidative metabolism enzymes, pyruvate dehydrogenase, aldehyde dehydrogenase and acetyl-CoA synthetase, decreased.     

Intensive rotifer cultures using chemostats

Continuous production of the rotifer Brachionus plicatilis rotundiformis (S-type) in an intensive chemostat culture system has been investigated. The production dynamics of rotifers in relation to different flow rates and feed regimes show that the growth rate and production depends on the type of algal feed and flow rate utilized in the culture system. It was possible to achieve a mean production of up to 318.84 × 10⁶ rotifers m^–3 d^–1 at a flow rate of 6 1 h^–1 in 100 1 chemostats and up to 261.21 × 10⁶ rotifers m^–3 d^–1 at a flow rate of 40 1 h –1 while using 1 m³ capacity rotifer chemostats as production units. The 3 fatty acid composition of rotifers while using Chlorella and Nannochloropsis in the culture system has been described. The results of this investigation show that the rotifer productivity in the continuous culture system is considerably higher than in any of the conventional culture systems described to date for aquacultural purposes.This research was financed by the Kuwait Foundation for the Advancement of Sciences (KFAS), Kuwait, under a contract research project code 86-04-02.     

Anaerobic degradation of p-cresol in batch and continuous cultures by a denitrifying bacterial consortium

Summary The anaerobic degradation of p-cresol under denitrifying conditions by a bacterial consortium was studied in batch and continuous cultures. Concentrations up to 3 mm were degraded within 5–6 days with 4-hydroxybenzyl alcohol, 4-hydroxybenzaldehyde and 4-hydroxybenzoate as intermediates. Steady states could be maintained at only one dilution rate, D=0.04 h^–1. A further increase in the dilution rate to 0.0 8 h^–1 resulted in culture wash-out. An estimation of the Saturation constant was made (<1 mg/l), taking the maximum specific growth rate as 0.045 h^–1, thus yielding a value of 0.125 mg p-cresol/l. Correspondence to: N. Khoury     

Volume flows and pressure changes during an action potential in cells ofChara australis

Summary Methods have been used for monitoring either volume flows or pressure changes, simultaneously with membrane potentials, in giant algal cells ofChara australis during an action potential. The volume flows were measured from the movement of a mercury bead in a capillary tube recorded by a photo-transducer. The pressure changes were measured by monitoring the deflection of a thin wedge, resting transversely across a cell, and using the same photo-transducer, the deflection of the wedge being directly related to the cell's turgor pressure. The average maximum rate of volume flow per unit area during an action potential was 0.88±0.11 nliter·sec^–1·cm^–2 in the direction of an outflow from the cell (total volume outflow being about 3 nliter·cm^–2 per action potential). Similarly, the maximum rate of change of pressure was 19.6±3.8×10^–3 atm·sec^–1 (peak change being 19.3±2.9×10^–3 atm equivalent to 14.7±2.2 mm Hg). The volume flow and pressure changes followed the vacuolar potential quite closely, the peak rate of volume flow lagging behind the peak of the action potential by 0.17±0.08 sec and the peak rate of pressure change leading it by 0.09±0.07 sec.     

Identification of two key genes controlling chill haze stability of beer in barley (Hordeum vulgare L)

Background

In bright beer, haze formation is a serious quality problem, degrading beer quality and reducing its shelf life. The quality of barley (Hordeum vulgare L) malt, as the main raw material for beer brewing, largely affects the colloidal stability of beer.

Results

In this study, the genetic mechanism of the factors affecting beer haze stability in barley was studied. Quantitative trait loci (QTL) analysis of alcohol chill haze (ACH) in beer was carried out using a Franklin/Yerong double haploid (DH) population. One QTL, named as qACH, was detected for ACH, and it was located on the position of about 108 cM in chromosome 4H and can explain about 20 % of the phenotypic variation. Two key haze active proteins, BATI-CMb and BATI-CMd were identified by proteomics analysis. Bioinformatics analysis showed that BATI-CMb and BATI-CMd had the same position as qACH in the chromosome. It may be deduced that BATI-CMb and BATI-CMd are candidate genes for qACH, controlling colloidal stability of beer. Polymorphism comparison between Yerong and Franklin in the nucleotide and amino acid sequence of BATI-CMb and BATI-CMd detected the corresponding gene specific markers, which could be used in marker-assisted selection for malt barley breeding.

Conclusions

We identified a novel QTL, qACH controlling chill haze of beer, and two key haze active proteins, BATI-CMb and BATI-CMd. And further analysis showed that BATI-CMb and BATI-CMd might be the candidate genes associated with beer chill haze.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1683-1) contains supplementary material, which is available to authorized users.     

Extraction of squalene from yeast by supercritical carbon dioxide

Squalene produced under anaerobic conditions, by a strain of Torulaspora delbrueckii was extracted from the biomass using supercritical carbon dioxide. Minimum use of solvent, lower time of isolation and a higher selectivity of extraction merit use of supercritical fluid extraction (SFE) technique over solvent extraction of squalene, as optimized and reported previously. A maximum squalene yield of 11.12 g g^–1 (dry weight) of yeast cells was obtained at a temperature of 60 °C and pressure of 250–255 bar at a constant flow rate of 0.2l min^–1 of carbon dioxide. Lyophilization prior to SFE increased the squalene yield to 430.52 g g^–1 dry weight of yeast cells, an amount that is far greater than that obtained by (2:1) chloroform–methanol solvent extraction.     

Factors which influence the sedimentation characteristics of Torulopsis glabrata after growth in a recirculation system

Summary Some environmental affects on cell aggregation described in the literature are briefly summarized. By means of a biomass recirculation culture (Contact system), using the yeast Torulopsis glabrata, the aggregation behavior of cells in static and in dynamic test systems is described. Sedimentation times required to obtain 50 g · l^–1 yeast dry matter in static systems were always higher than in dynamic ones.In addition to, influencing the biomass yield, the specific growth rate of the yeast also affected cell aggregation. The specific growth rate and therefore the aggregation could be regulated by the biomass recirculation rate as well as by the sedimenter volume.Abbreviations f_o Overflow flow rate (l·h^–1) - f_R Recycle flow rate (l·h^–1) - f_t0t Total flow rate through the fermenter (l·h^–1) - g Gram - h Hour - D_R Fermenter dilution rate due to recycle (h^–1) - D_S Fermeter dilution rate due to substrate (h^–1) - D_tot Total fermenter dilution rate (h^–1) - l Liter - Specific growth rate (h^–1) - P_F Fermenter productivity (g·l^–1·h^–1) - P_FS Overall productivity (g·l^–1·h^–1) - R_pM Rates per minute - RS Residual sugar content in the effluent with respect to the substrate concentration (%) - Y Yield of biomass with respect to sugar concentration (%) - Sed 50 Sedimentation time to reach a YDM of 50 g·l^–1 (min) - V Volume (l) - V_F Fermenter volume (l) - V_Sed Sedimenter volume (l) - VVM Volumes per volume and minute - X_F YDM in the fermenter (g·l^–1) - X_F YDM in the recycle (g·l^–1) - X_S Yeast dry matter due to substrate concentration (g·l^–1) - YDM Yeast dry matter (g·l^–1)