M. El Soda

Acceleration of cheese ripening with liposome-entrapped proteinase

SummaryRulactine, a proteinase used for the acceleration of cheese ripening, was entrapped in three types of liposomes and these were added to Saint-Paulin cheese type manufacturing milk. Enzyme entrapment rates ranged from 3 to 9% according to the type of liposomes and liposome retention rates in cheese curd from 35 to 65%. An electrophoretic study of protein breakdown in the cheeses gave correlative data.

Microfluidized liposomes for the acceleration of cheese ripening

Abstract Multilamellar (MLV), multilamellar-microfluidized (MLV-MF), microfluidized (MF) and dehydrated-rehydrated (DRV) liposomes were prepared and characterized. The mean diameters were 1360, 170, 110 and 1540 nm and the encapsulation efficiencies were 10, 14, 12 and 13% for MLV, MLV-MF, MF and DRV liposomes, respectively. Free and encapsulated trypsin in microfluidized liposomes were compared in one test trial for the acceleration of Cheddar cheese ripening. TCA soluble nitrogen data indicated an increase in protein degradation in both the free and entrapped enzyme cheese, compared to the control cheese. However, the action of the encapsulated trypsin was progressive as compared to the free enzyme treated cheese. Rheological measurements showed fracturability, firmness and cohesiveness defects in free enzyme cheese. Fracturability and firmness of the entrapped enzyme cheese were lower than the control cheese but cohesion was similar. This study indicates that the Microfluidizer™ can be successfully used for large scale production of liposomes without dissolving the lipids in organic solvents. The MF liposomes obtained could then be used for the acceleration of cheese ripening.

Autolytic properties and aminopeptidase activities of lactic acid bacteria

Abstract The ability of nine lactic acid bacteria ( Lactococcus , Lactobacillus and Pediococcus ) with potential in cheese maturation to lysis under various physicochemical conditions was studied. The lysis efficiency under various pH and NaCl concentrations was evaluated by measuring aminopeptidase and dipeptidylaminopeptidase activity released in cell supernatant. Two Lb. casei (L2A, 137) exhibited high peptidase activities and were more sensitive to lysis. Our results indicate clearly that autolysis is optimal with Lactobacillus casei casei (L2A) and Lactobacillus casei pseudoplantarum (137) at pH5.2 with 2–4% NaCl concentration and at 30 °C.

Microencapsulated enzyme systems for the acceleration of cheese ripening

Enzymes and substrates encapsulated in either milkfat-coated microcapsules or liposomes have been investigated for potential use as agents to accelerate cheese ripening. Milkfat-coated microcapsules have been used to efficiently encapsulate cell-free extracts, viable cells, purified enzymes, and spores. Encapsulation efficiency was dependent on the conditions used during capsule production. Addition of these microcapsules to cheese has resulted in increased levels of flavour compounds such as diacetyl, acetoin, methanethiol, and methyl ketones, compared to levels in control cheeses. Limitations due to cofactors have been overcome by co-encapsulating enzymes which recycle needed cofactors. Liposomes have been used to carry cell-free extracts and enzymes into cheese. Trials with different types of liposomes revealed that enzymes could be entrapped more efficiently and liposomes retained in the cheese curd better when multilammelar vesicles rather than small unilamellar vesicles or reverse phase evaporation vesicles were used. The stability of liposome preparations was found to be adversely affected by increased pH, temperature, and sodium chloride concentrations, as well as by negative surface charge. Encapsulation efficiency was found to increase by using a dehydration-rehydration procedure for liposome preparation. Temperature sensitive liposomes were investigates as a means of obtaining controlled release of the enzymes into the cheese.

Lipolytic activity of cheese related microorganisms and its impact on cheese flavour

Abstract Fat hydrolysis is one of the major biochemical events that takes place during cheese ripening. The action of carboxylesterases on the esters in the aqueous phase of the cheese probably also contributes to the formation of flavour compounds in cheese. The role of lactic acid bacteria and other cheese related microorganisms in fat degradation, the characterization and specificities of their lipases and esterases, as well as the influence of the growth conditions of the cells on enzyme production are discussed. Trends in future research have also been considered.

Autolysis of lactic acid bacteria: Impact on flavour development in cheese

Abstract Early autolysis of cheese related microorganisms in cheese should potentially release the intracellular enzymes into the cheese matrix. Such an event should lead to a more rapid development of flavour in cheese. The autolytic properties of several cheese related bacteria as well as the different factors affecting the process are discussed. Enzyme release during cell autolysis is also considered.

Cell-wall associated peptide hydrolase and esterase activities in several cheese-related bacteria

Cell-wall associated proteinases, peptidases and esterases of Lactobacillus helveticus, Pediococcus sp., Leuconostoc mesenteroides ssp. mesenteroides, Brevibacterium linens, Propionibacterium acidipropionici and Bifidobacterium infantis were assayed. Proteolytic activity was measured using C14-labelled casein. Aminopeptidase, dipeptidase and esterase activities were measured using chromogenic substrates. All the cheese related bacteria tested exhibited cell-wall proteinase activities with highest being detected in Leuconostoc mesenteroides ssp. mesenteroides, Pediococcus sp. and Brevibacterium linens. Lactobacillus helveticus and Brevibacterium linens exhibited greater cell-wall esterase and aminopeptidase activities, but little of the total activities of these enzymes were associated with the cell envelope. Greater properties of total cellular dipeptidase activities were associated with the cell envelope, with Lactobacillus helveticus and Brevibacterium linens exhibiting highest specific activities. The optimum pH of the crude cell-wall proteinase of different strains ranged from pH 5·5 to 8·0 while the optimum temperature ranged from 20 to 40°C. The impact of proteinase inhibitors tested differed between species.

The cell-bound proteinase system of Lactobacillus casei — purification and characterization

The cell-wall crude extract from Lactobacillus casei NCDO 151 was partially purified by DEAE-Sephacel chromatography. Three active fractions were eluted. Two major peaks (eluted with 0.05 M and 0.27 M phosphate buffer) were further investigated. Peak I represented enzymatic activity with an optimum temperature of 40 degrees C, an optimum pH of 7.0 and was strongly inhibited by the serine proteinase inhibitor phenylmethylsulfonylfluoride. Peak II represented an enzymatic activity with an optimum temperature of 45 degrees C, an optimum pH of 7.5 and was totally inhibited by p-hydroxymercuribenzoate. None of the enzymes was affected by the metal chelator ethylenediaminetetraacetic acid at a concentration up to 1 x 10(-2).

Purification of X-prolyl dipeptidyl aminopeptidase from Lactobacillus casei subspecies

Prolyl dipeptidylaminopeptidases from two subspecies of Lactobacillus casei were purified and biochemically characterized. L. casei ssp. casei UL21 (a debittering strain) and L. casei ssp. rhamnosus UL26 (a non-debittering strain) were the source bacteria for this study. Purification of the enzymes from both the sources was effected by a gel filtration step through Sephacryl S-300 followed by ion-exchange chromatography through DEAE Sephacel. This rendered an electrophoretically homogeneous enzyme preparation. The purified enzymes from both the sources showed similar temperature optimum (45 degrees C) and pH optimum (7.0). Their activity profiles on various substrates and the nature of inhibition by different inhibitors were also found to be similar, indicating that this enzyme is perhaps not significantly involved in the debittering process during the maturation of cheese.

Influence of microfiltration and adjunct culture on quality of Domiati cheese

The effects of microfiltration and pasteurization processes on proteolysis, lipolysis, and flavor development in Domiati cheese during 2 mo of pickling were studied. Cultures of starter lactic acid bacteria isolated from Egyptian dairy products were evaluated in experimental Domiati cheese for flavor development capabilities. In the first trial, raw skim milk was microfiltered and then the protein:fat ratio was standardized using pasteurized cream. Pasteurized milk with same protein:fat ratio was also used in the second trial. The chemical composition of cheeses seemed to be affected by milk treatment-microfiltration or pasteurization-rather than by the culture types. The moisture content was higher and the pH was lower in pasteurized milk cheeses than in microfiltered milk cheeses at d 1 of manufacture. Chemical composition of experimental cheeses was within the legal limits for Domiati cheese in Egypt. Proteolysis and lipolysis during cheese pickling were lower in microfiltered milk cheeses compared with pasteurized milk cheeses. Highly significant variations in free amino acids, free fatty acids, and sensory evaluation were found among the cultures used in Domiati cheesemaking. The cheese made using adjunct culture containing Lactobacillus delbrueckii ssp. lactis, Lactobacillus paracasei ssp. paracasei, Lactobacillus casei, Lactobacillus plantarum, and Enterococcus faecium received high scores in flavor acceptability. Cheeses made from microfiltered milk received a higher score in body and texture compared with cheeses made from pasteurized milk.