25 KB139 KB24 KB199820252000Golič Grdadolnik, SimonaLegiša, Matic12 straništevilka:3letnik:47str. 389-400ISSN:1318-0207COBISSID:2183706URN:URN:NBN:SI:doc-7P56HO35enSlovensko kemijsko društvoActa chimica slovenicanitaste gliveNMR spektroskopijapHImproved method for intracellular pH determination in filamentous fungi|Until now, it was difficult to accurately determine the intracellular pH of filamentous fungi growing in large bioreactors. In order not to change the pHsubi values during the preparation of samples and sup(31)P-NMR measurements, a new technique was developed that quenched the metabolism immediately after taking the cells out of the growing environment. It has been done by immersing the mycellium in cold buffered methanol at -40 supoC. The temperature of the samples was maintained low even during the accumulation of scans in the NMR magnet. Since frozen cells were surrounded by liquid buffered methanol, sufficient motion of molecules containing phosphorous was possible to enable the formation of high-resolution NMR spectra. The method has proven to be reliable enough for monitoring the changes of apparent intracellular pH that occur during hypoxia in a fermenterDo sedaj je bilo težko natančno določiti intracelularni pH pri filamentoznih glivah, ki so rastle v velikih bioreaktorjih. Da ne bi prišlo do spremembe pHspodaji vrednosti med pripravo vzorcev, oziroma med samo meritvijo Pzgoraj(31)jeder v NMR magnetu, smo razvili novo metodo, kjer smo metabolizem ustavili neposredno po tem, ko smo celice vzeli iz gojišča, in sicer tako, da smo jih potopili v ohlajen metanol pri -40 zgorajoC. Nizko temperaturo smo vzdrževali tudi kasneje med snemanjem v NMR magnetu. Ker so bile yamrznjene celice obdane s tekočim metanolom, so se molekule s fosforjem gibale v zadostni meri, da je lahko nastal NMR spekter visoke ločljivosti. Z uporabo nove metode smo uspeli zasledovati spreminjanje pH vrednosti v celicah med hipoksijo, ki smo jo povzročili s prekinjenim prezračevanjem v fermentorjuTEXTznanstveno časopisjejournalsSlovenian National E-content AggregatorNational and University Library of SloveniaSlovensko kemijsko društvo