Uloga Paecilomyces lilacinus (Thom) Samson i drugih vrsta gljiva u biodegradaciji ohratoksina A

Nine isolates of fungi of genera Aspergillus, Fusarium, Paecilomyces and Penicillium were cultured on the modified Vogel's medium with the addition of crude ochratoxin A (OTA) extract. This crude OTA extract was derived from a natural solid substrate on which Aspergillus ochraceus strain CBS 108.08 was cultivated. OTA was isolated, partially purified, dried by evaporating and dissolved in ethanol (1 mg ml-1), and added to the test medium up to the final concentration of 10 μg ml-1. The presence of OTA residues was determined after 7 and 14 day cultivation of fungi in the test medium at 27±1°C. The Paecilomyces lilacinus isolate (Inf. 2/A), which completely degraded OTA (150 μg) after only seven days, was selected for further studies. Wet sterile rice grains (50 g + 25 ml distilled water) were inoculated with individual isolates of fungi A. ochraceus (strain CBS 108.08) and P. lilacinus (isolate Inf. 2/A), and with their combination. In the case of P. lilacinus monoculture, 0.9 mg of crude OTA was also added into cultivation substrate. Each test was done in three replications. After the four week cultivation of individual and combined fungi at 27±1°C, inoculated rice grains were dried to the constant weight and pulverized. OTA was determined in these samples by the application of standard TLC method for fodder analysis. OTA in the amount of 61.310 μg kg-1 dry matter (DM) was determined only in the samples inoculated with a producer of ochratoxin A (A. ochraceus, strain CBS 108.08). On the other hand, a much smaller amount of OTA (80 μg kg-1 DM) was detected in samples inoculated with combined cultures of A. ochraceus and P. lilacinus isolates. Gained results indicate that P. lilacinus degraded, on average, 99.8% of OTA. After four week cultivation, the same fungal isolate in the samples of wet sterile rice kernels with the addition of 0.9 mg of crude OTA, completely degraded added crude OTA ( lt 8 μg kg-1).Devet izolata gljiva iz rodova Aspergillus, Fusarium, Paecilomyces i Penicillium gajeno je na modifikovanoj Vogelovoj podlozi sa dodatkom sirovog ekstrakta ohratoksina A (OTA). Sirovi ekstrakt OTA je dobijen iz čvrstog prirodnog supstrata na kojem je gajen soj Aspergillus ochraceus CBS 108.08. Izolovan i delimično prečišćen OTA, uparen do suvog ostatka i rastvoren u etanolu (1 mg ml-1), dodat je u test podlogu do finalne koncentracije 10 μg ml-1. Nakon sedam i 14 dana gajenja kultura gljiva u test podlozi na 27 ± 1°C de terminisano je prisustvo rezidua OTA primenom modifikovane metode Filtenborg-a i sar. (1983). Od devet testiranih izolata za dalja ispitivanja je odabran izolat Paecilomyces lilacinus (Inf. 2/A), koji je već posle sedam dana u potpunosti razgradio inicijalnu količinu OTA (150 μg). U drugom delu eksperimenta vlažno sterilno zrno pirinča (50 g + 25 ml destilovane vode) zasejano je sa pojedinačnim izolatima A. ochraceus (CBS 108.08) i P. lilacinus (Inf. 2-A), kao i kombinacijom oba izolata. U slučaju mono-kulture P. lilacinus u podlogu je dodat i sirovi OTA (0,9 mg). Svaki od testova je urađen u 3 ponavljanja. Nakon četiri nedelje gajenja monokultura i mešanih kul tura gljiva na 27±1°C, inokulisana zrna su osušena do konstantne težine i sa mlevena do finog praha. U ovim uzorcima izvršena je determinacija OTA primenom standardne metode tankoslojne hromatografije za analizu stočne hrane. U uzorcima koji su bili zasejani samo sa producentom OTA (A. ochraceus, soj CBS 108.08) detektovan je OTA u prosečnoj količini od 61.310 μg kg-1 suvog ostatka. U uzorcima koji su bili zasejani kombinovanim kulturama izolata A. ochraceus i P. lilacinus utvrđena je znatno manja prosečna količina OTA (80 μg kg-1). Ovi rezultati ukazuju da je izolat P. lilacinus razgradio prosečno 99,8% OTA prisutnog u podlozi za kultivaciju. U uzorcima vlažnog sterilnog zrna pirinča sa dodatkom 0,9 mg sirovog OTA isti gljivični izolat je posle četiri nedelje kultivacije kompletno biorazgradio dodat sirovi OTA ( lt 8 μg kg-1)

In vitro degradacija diacetoksiscirpenola i T-2 toksina posredstvom izolata Mucor racemosus fresen. f. racemosus

Under controlled in vitro conditions the capacity of the Mucor racemosus f. racemosus 1215/09 isolate to degrade type A trichothecenes (diacetoxyscirpenol - DAS and T-2 toxin) was observed in the liquid nutritive medium. According to previously performed experiments it was proved that the selected isolate, originating from sunflower meal, had the ability to degrade these fusariotoxins when growing on the modified Vogel's agar supplemented with crude extracts of DAS and T-2 toxin. In order to determine biodegradation of fusariotoxins, the liquid nutritive medium - SPY (5% sucrose + 0.1% peptone + 0.1% yeast extract, pH 6.2) was simultaneously inoculated with the isolate M. racemosus f. racemosus 1215/09 and: a) Fusarium semitectum SL-B (DAS producer) or b) F. sporotrichioides R-2301 (T-2 toxin producer). The SPY media, inoculated with single fungal isolates, were used as a control of toxin biosynthesis. The cultures were incubated at room temperature (21-26ºC) on the rotary shaker (175 rpm). After the 3-5-day incubation, the filtration of liquid cultures and the extraction of fusariotoxins from filtrates with ethyl-acetate were performed. Determinations of DAS and T-2 toxin were done by thin layer chromatography using silica gel G. Depending on the incubation duration, M. racemosus f. racemosus in the mixed culture with F. semitectum degraded from 90.0 to 99.97% of DAS present in the medium (40,000- 120,000 µg l-1), while in the mixed culture with F. sporotrichioides it degraded from 95.0 to 96.7% of T-2 toxin present in the medium (240,000 µg l-1). Sterile filtrates of mixed cultures and single culture of M. racemosus f. racemosus, obtained by passing liquid cultures through the 0.45-µm membrane filter and added to the SPY medium, did not affect degradation of type A trichothecenes that had been biosynthesised by isolates F. semitectum SL-B and F. sporotrichioides R-2301 in the liquid medium.U kontrolisanim in vitro uslovima proučavana je sposobnost izolata Mucor racemosus f. racemosus 1215/09 da degraduje trihotecene tipa A (diacetoksiscirpenol - DAS i T-2 toksin) u tečnoj hranljivoj podlozi. Prethodnim eksperimentima je dokazano da odabrani izolat, poreklom sa suncokretove sačme, poseduje sposobnost razgradnje navedenih fuzariotoksina, koji su kao sirovi ekstrakti dodati u modifikovanu Vogelovu podlogu. U cilju utvrđivanja biodegradacije fuzariotoksina tečna hranljiva podloga SPK (5% saharoza + 0,1% pepton + 0,1% ekstrakt kvasca, pH 6,2) je zasejana u isto vreme izolatom M. racemosus f. racemosus 1215/09 i: a) Fusarium semitectum SL-B (proizvođač DAS-a) ili b) F. sporotrichioides R-2301 (proizvođač T-2 toksina). Kao kontrola biosinteze toksina korišćena je SPK podloga inokulisana pojedinačnim izolatima gljiva. Kulture su inkubirane na rotacionoj tresilici (175 o/min) tokom 3-5 dana na sobnoj temperaturi (21-26ºC). Nakon 3 do 5 dana inkubacije vršeno je filtriranje tečnih kultura i ekstrakcija fuzariotoksina iz filtrata etil-acetatom. Determinacija DAS-a i T-2 toksina je rađena tankoslojnom hromatografijom na silika gelu G. Zavisno od dužine inkubacije, M. racemosus f. racemosus je u združenoj kulturi sa F. semitectum degradovala 90,0-99,97% DAS-a prisutnog u podlozi (40.000-120.000 µg l-1), dok je u združenoj kulturi sa F. sporotrichioides razgradila 95,0-96,7% T-2 toksina prisutnog u podlozi (240.000 µg l-1). Sterilni filtrati mešanih kultura i pojedinačne kulture M. racemosus f. racemosus, dobijeni propuštanjem tečnih kultura kroz 0,45 µm membranski filter i dodati SPK podlozi, nisu uticali na razgradnju trihotecena tipa A koje su biosintetisali izolati F. semitectum SL-B i F. sporotrichioides R-2301 u tečnoj podlozi

Uticaj peletiranja na mikrobiološku i mikotoksikološku ispravnost krmnih smeša sa dodatkom bentonita

Influence of pelleting calf feed mixtures supplemented with bentoni on microbiological and mycotoxicological properties was investigated. Microbiological and mycotoxicological quality was investigated at the production day (day 0) and after 45 days of storage. Total count of microorganisms in the pelleted mixture, at the day 0 (280.000/g), was several times lower than in the powdered mixture (2.000.000/g). Similar results were obtained at day 45 when the total number of microorganisms in the pelleted mixture was 270.000/g and 1.800.000/g in the powdered mixture. Number of yeasts and molds at the production day in the pelleted mixture was 650/g, and in the powdered mixture it was 27.000/g. Similar results were obtained 45 days later when the number of yeasts and molds in the pelleted mixture was 540/g, and 16.000/g in the powdered mixture. There were 6 species identified in the pelleted mixture, and 9 species in the powdered mixture at the day of production. Similar mold species ratio in the pelleted (11) and powdered mixture (13) was found at day 45. In the examined samples representatives of Fusarium genus - F. subglutinans i F. verticillioides dominated. Number of sultite-redzcing clostridia in the mixtures, in both observed periods, was similar (below 1000/g of sample). By mycotoxicological analysis of mixtures at the production day, only trichotecene (T-2 toxin) presence was found in amount of 0,337 mg/kg. The applied technological procedure of pelleting with bentonite supplement, had positive influence on the improvement of microbiological and toxicological properties of mixture.U ogledu je ispitivan uticaj peletiranja krmnih smeša za telad sa dodatkom bentonita na mikrobiološku i mikotoksikološku ispravnost smeša. Brašnasta i peletirana krmna smeša za telad su proizvedene po istoj recepturi. Mikrobiološka i mikotoksikološka ispravnost smeša ispitana je na dan proizvodnje (0-ti dan) i posle 45 dana lagerovanja. Ukupan broj mikroorganizama u peletiranoj smeši, na dan proizvodnje (280.000/g) bio je višestruko manji od broja u brašnastoj smeši (2.000.000/g). Slično je bilo 45 dana kasnije, kada je ukupan broj mikroorganizama u peletiranoj smeši iznosio 270.000/g, odnosno 1.800.000/g u brašnastoj smeši. Broj kvasaca i plesni na dan proizvodnje u peletiranoj smeši bio je 650/g, a u brašnastoj 27.000/g. Slični rezultati utvrđeni su 45 dana kasnije, kada je broj kvasaca i plesni u peletiranoj smeši iznosio 540/g, a u brašnastoj 16.000/g. U peletiranoj smeši na dan proizvodnje identifikovano je 6 vrsta, a u brašnastoj 9 vrsta plesni. Sličan odnos vrsta plesni u peletiranoj (11) i brašnastoj (13) utvrđen je i 45 dana kasnije. U ispitanim uzorcima su dominirali predstavnici roda Fusarium - F. subglutinans i F. verticillioides. Broj sulfitoredukujućih klostridija u smešama, u oba termina kontrole, bio je sličan, odnosno ispod 1000/g uzorka. Ostale vrste patogenih bakterija nisu identifikovane. Mikotoksikološkom analizom smeša na dan proizvodnje utvrđeno je jedino prisustvo trihotecena (T-2 toksin) u količini od 0,337 mg/kg smeše. Primenjeni tehnološki postupak peletiranja, uz dodatak bentonita kao vezivnog sredstva, imao je pozitivan uticaj na poboljšanje mikrobiološke i toksikološke ispravnosti ispitivanih krmnih smeša

Synthesis and characterization of high-pressure and high-temperature sphene (CaTiSiO5)

Sphene (CaTiSiO5), a calcium titanosilicate ceramic has been prepared from a powder mixture of CaCO3, TiO2 and SiO2 using vibro-milling for homogenization and activation of precursors. During the high-pressure and high-temperature synthesis (HPS) process at 4 GPa and 1,200 A degrees C, sphene undergoes into phase transition, from room-temperature phase P2(1) /a to high-temperature phase A2/a. Evidence of that structural phase transition is given in this paper using infrared, Raman spectroscopy and X-ray powder diffraction. Rietveld refinement was employed to get the structural information of the synthesized powder. The most important structural change due to phase transition, the disappearance of the characteristic out-of-center distortion of the Ti atom and moving to the center of octahedra, was confirmed. HPS is an effective method for producing full-dense ceramics without any additives. Reduction of particle size occurred during high-pressure compaction. Microstructure and particle size of both phases were analyzed by scanning electron microscopy

Influence of pelleting on microbiological and mycotoxical correctness of feed mixtures with bentonite supplement

Influence of pelleting calf feed mixtures supplemented with bentoni on microbiological and mycotoxicological properties was investigated. Microbiological and mycotoxicological quality was investigated at the production day (day 0) and after 45 days of storage. Total count of microorganisms in the pelleted mixture, at the day 0 (280.000/g), was several times lower than in the powdered mixture (2.000.000/g). Similar results were obtained at day 45 when the total number of microorganisms in the pelleted mixture was 270.000/g and 1.800.000/g in the powdered mixture. Number of yeasts and molds at the production day in the pelleted mixture was 650/g, and in the powdered mixture it was 27.000/g. Similar results were obtained 45 days later when the number of yeasts and molds in the pelleted mixture was 540/g, and 16.000/g in the powdered mixture. There were 6 species identified in the pelleted mixture, and 9 species in the powdered mixture at the day of production. Similar mold species ratio in the pelleted (11) and powdered mixture (13) was found at day 45. In the examined samples representatives of Fusarium genus - F. subglutinans i F. verticillioides dominated. Number of sultite-redzcing clostridia in the mixtures, in both observed periods, was similar (below 1000/g of sample). By mycotoxicological analysis of mixtures at the production day, only trichotecene (T-2 toxin) presence was found in amount of 0,337 mg/kg. The applied technological procedure of pelleting with bentonite supplement, had positive influence on the improvement of microbiological and toxicological properties of mixture

The role of paecilomyces lilacinus (thom) samson and other fungal species in biodegradation of ochratoxin a

Nine isolates of fungi of genera Aspergillus, Fusarium, Paecilomyces and Penicillium were cultured on the modified Vogel’s medium with the addition of crude ochratoxin A (OTA) extract. This crude OTA extract was derived from a natural solid substrate on which Aspergillus ochraceus strain CBS 108.08 was cultivated. OTA was isolated, partially purified, dried by evaporating and dissolved in ethanol (1 mg ml-1), and added to the test medium up to the final concentration of 10 μg ml-1. The presence of OTA residues was determined after 7 and 14 day cultivation of fungi in the test medium at 27±1°C. The Paecilomyces lilacinus isolate (Inf. 2/A), which completely degraded OTA (150 μg) after only seven days, was selected for further studies. Wet sterile rice grains (50 g + 25 ml distilled water) were inoculated with individual isolates of fungi A. ochraceus (strain CBS 108.08) and P. lilacinus (isolate Inf. 2/A), and with their combination. In the case of P. lilacinus monoculture, 0.9 mg of crude OTA was also added into cultivation substrate. Each test was done in three replications. After the four week cultivation of individual and combined fungi at 27±1°C, inoculated rice grains were dried to the constant weight and pulverized. OTA was determined in these samples by the application of standard TLC method for fodder analysis. OTA in the amount of 61.310 μg kg-1 dry matter (DM) was determined only in the samples inoculated with a producer of ochratoxin A (A. ochraceus, strain CBS 108.08). On the other hand, a much smaller amount of OTA (80 μg kg-1 DM) was detected in samples inoculated with combined cultures of A. ochraceus and P. lilacinus isolates. Gained results indicate that P. lilacinus degraded, on average, 99.8% of OTA. After four week cultivation, the same fungal isolate in the samples of wet sterile rice kernels with the addition of 0.9 mg of crude OTA, completely degraded added crude OTA (<8 μg kg-1)

Microencapsulation of anthocyanin-rich black soybean coat extract by spray drying using maltodextrin, gum Arabic and skimmed milk powder

Black soybean coat is insufficiently valorised food production waste rich in anthocyanins. The goal of the study was to examine physicochemical properties of spray dried extract of black soybean coat in regard to carrier materials: maltodextrin, gum Arabic, and skimmed milk powder. Maltodextrin and gum Arabic-based microparticles were spherical and non-porous while skimmed milk powder-based were irregularly shaped. Low water activity of microparticles (0.31-0.33), good powders characteristics, high solubility (80.3-94.3%) and encapsulation yields (63.7-77.0%) were determined. All microparticles exhibited significant antioxidant capacity (243-386 mu molTE/g), good colour stability after three months of storage and antimicrobial activity. High content of total anthocyanins, with cyanidin-3-glucoside as predominant, were achieved. In vitro release of anthocyanins from microparticles was sustained, particularly from gum Arabic-based. These findings suggest that proposed simple eco-friendly extraction and microencapsulation procedures could serve as valuable tools for valorisation and conversion of black soybean coat into highly functional and stable food colourant