PREDICTION OF THE SAFETY OF GLAZED CONFECTIONERY PRODUCTS BY THE CONTENT OF LIPOLYTIC MICROORGANISMS

Гидролиз жиров лауринового типа, вызываемый липолитическими ферментами, может придавать глазированным кондитерским изделиям при хранении неприятный «мыльный» привкус, связанный с накоплением свободной лауриновой кислоты. Активность липазы в кондитерских изделиях может существенно изменяться в процессе хранения при микробиологической контаминации, поскольку микроорганизмы – бактерии, плесневые грибы и дрожжи являются активными продуцентами липаз. Проведен мониторинг качества глазурей, изготовленных на основе жиров лауринового типа, различных кондитерских изделий (кондитерских плиток; образцов глазури различных наименований, полученных от производителя, глазури, отделенной от мучных кондитерских изделий и корпусов конфет типа пралине, помадные, сбивные (всего 45 образцов)), в результате которого установлено, что содержание КМАФАнМ во всех образцах глазури составило не более 105 КОЕ/г продукта, дрожжей – менее 50 КОЕ/г, плесеней – 50–100 КОЕ/г. Для исследования влияния микробиологической обсемененности на изменение кислотности кондитерского изделия в процессе хранения были изготовлены модельные образцы глазури с высоким содержанием лауриновой кислоты и долей общего сухого остатка какао 15, 25 и 28%, инокулированные тест-культурами Subtilis Bacillus КВ2-1 и Aerophilus Bacillus 6-1 в количестве 10, 102, 103 и 104 КОЕ/г. На основе математической обработки экспериментальных данных построены графики зависимости кислотности от доли общего сухого остатка какао и обсемененности модельных образцов при хранении в течение 12 мес. Выявлена математическая зависимость, представленная в виде квадратичной функции, позволяющая прогнозировать сохранность глазированных кондитерских изделий. Экстремум функции показывает период хранения глазури, характеризующийся наибольшей скоростью образования свободных жирных кислот. Установлено, что при увеличении микробиологической обсемененности, а также при снижении массовой доли общего сухого остатка какао в какаосодержащем полуфабрикате период хранения, характеризующийся наибольшей скоростью образования свободных жирных кислот, сокращается. На основе полученных закономерностей возможно прогнозирование изменений органолептических показателей глазированных кондитерских изделий, а также кондитерских плиток и фигурных изделий, содержащих жиры лауринового типа, в процессе хранения. The hydrolysis of lauric-type fats caused by lipolytic enzymes can give glazed confectionery products an unpleasant «soapy» taste associated with the accumulation of free lauric acid during their storage. The activity of lipase in confectionery products can significantly change during storage during microbiological contamination, since microorganisms – bacteria, mold fungi and yeast are active lipase producers. Monitoring of the quality of glazes made on the basis of laurine-type fats, various confectionery products (confectionery tiles; glaze samples of various names received from the manufacturer, glaze separated from flour confectionery products and sweets with praline, fondant, whipped cases (45 samples in total)) conducted. According to its results, it was found that the content of CMAFAnM in all glaze samples was no more than 105 CFU/g of the product, yeast – less than 50 CFU/g, molds – 50–100 CFU/g. To study the effect of microbiological contamination on the change in the acidity of the confectionery product during storage, model glaze samples with a high content of lauric acid and a share of the total cocoa solids of 15, 25 and 28%, innoculated with test cultures of Subtilis Bacillus KV2-1 and Aerophilus Bacillus 6-1 in the amount of 10, 102, 103 and 104 CFU/g, were made. On the basis of mathematical processing of experimental data, graphs of the dependence of acidity on the proportion of the total cocoa solids and the contamination of model samples during storage for 12 months were constructed. A mathematical dependence, presented in the form of a quadratic function, which allows predicting the safety of glazed confectionery products, has been identified. The extremum of the function shows the storage period of the glaze, characterized by the highest rate of formation of free fatty acids. It was found that with an increase in microbiological contamination, as well as with a decrease in the mass fraction of the total cocoa solids in the cocoa-containing semi-finished product, the storage period characterized by the highest rate of formation of free fatty acids is reduced. On the basis of the obtained regularities, the prediction of changes in the organoleptic parameters of glazed confectionery products, as well as confectionery tiles and figured products containing lauric-type fats, during storage can be carried out.

Kinetic and Transesterification Properties of Lipase from Sprouted Melon (Cucumeropsis manni) Seeds

Crude lipase (acetone powder) was extracted from freshly sprouted melon seeds (Cucumeropsis manni). The activity, kinetic properties (effect of time, pH, and enzyme and substrate concentration, respectively) as well as the ability of the crude lipase to catalyze the production of methyl esters (biodiesel) were examined. The enzyme activity was determined using n-hexane as the solvent (1:2 v/w solvent: substrate ratio) and the transesterification product was analyzed by HPLC. A linear relationship was observed between reaction time and rate of lipolysis with the optimal activity at 2hr of incubation. Furthermore, the lipase was optimally active at acid pH 5 and lipolysis was achieved optimally when the amount of enzyme was 2.0g. Rate of lipolysis was observed to increase linearly at concentrations up to 5.0g of substrate above which a drop in the rate, with no apparent decrease in activity, was observed. The Km (6.25g) and Vmax (13.33%FFA/hr) were also determined. Analysis of the transesterification product yielded 0.61% alkyl ester, 0.81 %FFA, 93.17% TAG, 4.15% 1, 3-DAG and 1.26% 1, 2-DAG while transesterification efficiency was determined to be at 0.588%. Biodiesel (alkyl esters) prepared with the crude lipase was had a density of 0.872 g/mL while its cloud and pour points were 22°C and 12°C, respectively. The results from this research showed that an active lipase was isolated from sprouted melon seeds. However, the fuel properties of the biodiesel produced did not meet international transportation fuel standards. In order to be used industrially, better reaction conditions need to be established for the lipase.

Cold-Active Lipase-Based Biocatalysts for Silymarin Valorization through Biocatalytic Acylation of Silybin

Extremophilic biocatalysts represent an enhanced solution in various industrial applications. Integrating enzymes with high catalytic potential at low temperatures into production schemes such as cold-pressed silymarin processing not only brings value to the silymarin recovery from biomass residues, but also improves its solubility properties for biocatalytic modification. Therefore, a cold-active lipase-mediated biocatalytic system has been developed for silybin acylation with methyl fatty acid esters based on the extracellular protein fractions produced by the psychrophilic bacterial strain Psychrobacter SC65A.3 isolated from Scarisoara Ice Cave (Romania). The extracellular production of the lipase fraction was enhanced by 1% olive-oil-enriched culture media. Through multiple immobilization approaches of the cold-active putative lipases (using carbodiimide, aldehyde-hydrazine, or glutaraldehyde coupling), bio-composites (S1–5) with similar or even higher catalytic activity under cold-active conditions (25 °C) have been synthesized by covalent attachment to nano-/micro-sized magnetic or polymeric resin beads. Characterization methods (e.g., FTIR DRIFT, SEM, enzyme activity) strengthen the biocatalysts’ settlement and potential. Thus, the developed immobilized biocatalysts exhibited between 80 and 128% recovery of the catalytic activity for protein loading in the range 90–99% and this led to an immobilization yield up to 89%. The biocatalytic acylation performance reached a maximum of 67% silybin conversion with methyl decanoate acylating agent and nano-support immobilized lipase biocatalyst.

Efficient Biodiesel Production Catalyzed by Nanobioconjugate of Lipase from Pseudomonas fluorescens

The Amano lipase from Pseudomonas fluorescens (L-AK) was covalently immobilized on various carbon nanomaterials (functionalized single-walled carbon nanotubes and graphene oxide) and tested for biodiesel production. Using the most active lipase preparation (covalently immobilized L-AK on SwCNTNH2 derivatized with glycerol diglycidyl ether) under optimal conditions, quasi-complete conversion (>99%) of sunflower oil was obtained after only 4 h reaction time. Moreover, the biocatalyst maintained more than 99% of its initial activity in the batch system after multiple recycling experiments.