CS210222B1 - A method for producing digalacturonic acid by enzyme hydrolysis - Google Patents

A method for producing digalacturonic acid by enzyme hydrolysis Download PDF

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CS210222B1
CS210222B1 CS142580A CS142580A CS210222B1 CS 210222 B1 CS210222 B1 CS 210222B1 CS 142580 A CS142580 A CS 142580A CS 142580 A CS142580 A CS 142580A CS 210222 B1 CS210222 B1 CS 210222B1
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acid
digalacturonic
digalacturonic acid
producing
soluble
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CS142580A
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Czech (cs)
Slovak (sk)
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Kveta Heinrichova
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Kveta Heinrichova
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Abstract

Vynález sa týká spósobu přípravy kyseliny digalakturónovej enzýmovou hydrolýzou. Podstata vynálezu spočívá v tom, že se na kyselinorozpustnú polygalakturónovú kyselinu pósobí enzýmom endo-D-galakturonanázou pri teplote 30 až 40 °C a pH 4»2 až 5, pričom kyselina digalakturónová sa oddělí chromatograficky na koloně a efluent sa odpaří alebo vysuší mrazovou sublimáciou. Význam vynálezu spočívá v použití kyseliny digalakturónovej na testovanie róznych enzymatických preparátov používaných v konzervárskom, kožiarskom a textilnom priemysle.The invention relates to a method of preparing digalacturonic acid by enzymatic hydrolysis. The essence of the invention is that acid-soluble polygalacturonic acid is treated with the enzyme endo-D-galacturonanase at a temperature of 30 to 40 °C and pH 4»2 to 5, whereby digalacturonic acid is separated by column chromatography and the effluent is evaporated or dried by freeze sublimation. The significance of the invention lies in the use of digalacturonic acid for testing various enzymatic preparations used in the canning, leather and textile industries.

Description

Vynález sa týká spĎeobu přípravy kyaeliny digalakturonovej enzýmovou hydrolýzou.The present invention relates to a process for preparing kyaeline by digalacturon enzyme hydrolysis.

Kyaelina digalakturonová aa doteraz priemyeelne navyrába. Bola připravené v niekol'-kých laboratoriách ceatou chemickéj hydrolýzy (Hátanaka C., Ozawa J. J. Agr. Chem. Soc.JapanVol. 40 etr. 98 (1966); Rexová-Benková L., Chem. zvěsti 24 (1970), atr. 59; van Hon-denhoven F., Wittp. D., Visser J. Carbohyd, Res. 34, atr. 233 (1974)). Všetky popíaané po-stupy sú viacatupňové a značné sa odlišujú heterogenitou získaného produktu. Spoločným ry-aom týchto poatupov je, že vedú k malým výťažkom kyseliny digalakturonovej popři vysokomvýtažku vyěších oligogalakturo'nových kyselin. Získanie kyseliny digalakturonovej v čistomstave zo zmesi oligogalakturonových kyselin má mnohé nevýhody ako je pracnost, časová ná-ročnost a viacatupňové izolácie,Kyaelina digalacturone and up to now industryeelne. It has been prepared in several laboratories by chemical hydrolysis (Hatanaka C., Ozawa JJ Agr. Chem. Soc.JapanVol. 40 et al. 98 (1966); Rex-Benková L., Chem. 24 (1970), atr. 59, van Hon denhoven F., Wittp D., Visser J. Carbohyd, Res 34, atr 233 (1974). All of the processes described are multi-stage and considerable differ in the heterogeneity of the product obtained. The common feature of these processes is that they result in low yields of digalacturonic acid in addition to the high yield of the higher oligogalacturic acids. Obtaining digalacturonic acid in a purification of oligogalacturonic acid mixtures has many disadvantages, such as labor, time-consuming and multi-stage insulation,

Tieto nedostatky odstraňuje spdsob výroby kyseliny digalakturonovej enzýmovou hydro-lýzou, ktorého podstata spočívá v tom, že sa na kyselinorozpustnú polygalakturonovú kyse-linu pĎsobí enzýmom endo-D-galakturonanázou pri teplote 30 až 40 °C a pH 4,2 až 5, pričomkyselina digalakturonová sa oddělí chromatograficky na kolo'ne a efluent sa odpaří alebo vy-suší mrazovou sublimáciou. Výhodou tohto postupu je, že endo-D-galakturonáza sa nemusí izolovat v čistom stave.These drawbacks are eliminated by the process for the production of digalacturonic acid by enzymatic hydrolysis, wherein the acid-soluble polygalacturonic acid is treated with the enzyme endo-D-galacturonanase at 30-40 ° C and pH 4.2-5, wherein the digalacturonic acid they are separated by column chromatography and the effluent is evaporated or freeze-dried. The advantage of this procedure is that the endo-D-galacturonase need not be isolated in a pure state.

Na enzýmovú hydrolýzu sa mdže použit endo-D-galakturonanáza z komerčných preparátov pekto-lytických enzýmov (Pectinase, Rohament, Pectinex Ultra, Pekticín, Leozym a i.) bežne použí-vaných v konzervárenskom priemysle. Jedinou požiadavkou na použitý enzýmový preparát jenízký obsah exo-D-galakturonanázy. fialšou velkou přednostou uvedeného apčsobu je, že hyd-rolýza prebieha formou jednostupňovej reakcie a pře priebeh reakcie nie je potřebný eialšísubstrát alebo koenzým.For enzymatic hydrolysis, endo-D-galacturonanase can be used from commercial preparations of cytolytic enzymes (Pectinase, Rohament, Pectinex Ultra, Pecticin, Leozym and others) commonly used in the canning industry. The only requirement for the enzyme preparation used is the high exo-D-galacturonanase content. the more vivid advantage of said process is that the hydrolysis takes place in the form of a one-step reaction and that the reaction is not required to be a substitute or coenzyme.

Kyselina digalakturonová je látka dfiležitá z hlediska teoretického štúdia enzýmova pektínových látok a pre charakterizáciu v priemysle používaných pektolytických enzýmov. Příklad 1Digalacturonic acid is a substance that is relevant to the theoretical study of enzyme pectin substances and to the characterization of the pectolytic enzymes used in the industry. Example 1

Pri príprave substrátu (kyselinorozpustnej polygalakturónovej kyseliny) pre hydrolý-zu sa postupovalo podlá upraveného postupu Mc Creadi Seegmiller: Archiv of Biochim. andBiophys. 50, 440, 1954. 50 g pektínu sa za stálého miešania přidává do 2 1 1N kyselinysirovej. Po rozpuštění sa suspenzia zahrieva pri 100 °0 60 minút. Suspenzia sa chladía v kyselině nerozpustný podiel sa oddělí filtráciou. Supernatant sa zráža 2 litrami etyl-alkoholu a nechá stát 24 hodin. Vyzrážaná kyselinorozpustná polygalakturonová kyselina saoddělí centrifugováním při 10 000 g. Supernatant sa zráža štyrmi litrami 96 % etylalkoho-lu a postup sa opakuje ako v prvom případe. Obidva podiely vyzrážanej kyselinorozpustnejpolygalakturónovej kyseliny sa spoja, dvakrát premyjú 70 % etylalkoholom a nakoniec 1 krátacetónom. Preparát sa volné vysuší pri laboratornej teplote alebo sa rozpustí v destilova-nej vodě a vysuší mrazovou sublimáciou. Příklad 2 500 mg kyselinorozpustnej polygalakturonovej kyseliny o priemernom polymerizačnomstupni 10, připravenéj podlá příkladu 1, sa postupné za miešania rozpustí v 70 ml octano-The preparation of the substrate (acid-soluble polygalacturonic acid) for hydrolysis was carried out according to the modified Mc Creadi Seegmiller procedure: Archiv of Biochim. andBiophys. 50, 440, 1954. After dissolution, the suspension is heated at 100 ° C for 60 minutes. The suspension was cooled in an acid-insoluble manner by filtration. The supernatant is precipitated with 2 liters of ethyl alcohol and allowed to stand for 24 hours. The precipitated acid-soluble polygalacturonic acid is separated by centrifugation at 10,000 g. The supernatant is precipitated with four liters of 96% ethyl alcohol and the procedure repeated as in the first case. The two portions of the precipitated acid-soluble polygalacturonic acid are combined, washed twice with 70% ethyl alcohol and finally with 1-acetone. The preparation is freely dried at room temperature or dissolved in distilled water and freeze-dried. EXAMPLE 2 500 mg of an acid-soluble polygalacturonic acid having an average polymerization stage of 10 prepared according to Example 1 was dissolved in 70 ml of acetone, with stirring.

Claims (1)

PBEDMET VYNÁLEZUTHE PBEDMET OF THE INVENTION SpSsob výroby kyseliny digalakturonovej enzýmovou hydrolýzou vyznačujúci sa tým, že sa na kyselinorozpustnú polygalakturónovú kyselinu pdsobí enzýmom endo-D-galakturonanázou pri teplote 30 až 40 °C a pH 4,2 až 5, pričom kyselina digalekturonová sa oddělí chromatograficky na koloně a efluent sa odpaří alebo vysuší mrazovou sublimáciou.Process for the preparation of digalacturonic acid by enzymatic hydrolysis, characterized in that acid-soluble polygalacturonic acid is treated with the enzyme endo-D-galacturonanase at a temperature of 30 to 40 ° C and a pH of 4.2 to 5, wherein the digalecturonic acid is separated by column chromatography and the effluent is evaporated or freeze-dried.
CS142580A 1980-03-03 1980-03-03 A method for producing digalacturonic acid by enzyme hydrolysis CS210222B1 (en)

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