Microencapsulation of flavour-enhancing enzymes for acceleration of cheddar cheese ripening

Commercial flavour-enhancing enzymes were delivered in an encapsulated form to accelerate Cheddar cheese ripening. Polymers such as alginate, chitosan and k- Carrageenan were screened to be used as encapsulant material for microencapsulation of the commercial protease enzyme, Flavourzyme®. Alginate was found to be a suitable polymer for Flavourzyme encapsulation using the Inotech® encapsulator while _-Carrageenan and chitosan were too viscous for extrusion through the encapsulator nozzle. Gelling of alginate-Flavourzyme microcapsules in 0.1M CaCl2 resulted in poor encapsulation efficiency (ranging 17- 18% depending on the alginate concentration). Incorporation of Hi-Maize™ starch or pectin as filler materials into the alginate-Flavourzyme encapsulation matrix to increase encapsulation efficiency by minimising porosity also resulted in poor encapsulation efficiency. An alternative approach to the modification of the cationic gelling solution, by adding chitosan, significantly increased the encapsulation efficiency to 70-88% and produced mostly spherical capsules with an average diameter of 500_m. Encapsulation efficiency increased with an increase in chitosan concentration from 0.1 to 0.3% (w/v) in the cationic gelling solution of 0.1M CaCl2. Though gelling of alginate-Flavourzyme microcapsules in gelling solution of 0.1M CaCl2 containing 0.3% (w/v) chitosan resulted in higher encapsulation efficiency, a chitosan concentration of 0.1% (w/v) was chosen for further work as higher concentrations of chitosan in the gelling solution resulted in aggregation of capsules during formation. Gelling time of 10 min and alginate concentrations in the range 1.6 to 2.0% (w/v) were found to be optimal encapsulation parameters for Flavourzyme encapsulation while 2.0% (w/v) solution of trisodium citrate was found to be optimal for in vitro release of encapsulated enzymes for measurement of enzyme activity. Flavourzyme capsules stored frozen or freeze-dried were shelf stable for at least 10 weeks retaining about 80% of the initial enzyme activity as opposed to retention of 25-34% activity in air-dried capsules. Leakage of encapsulated Flavourzyme prepared from 1.6% (w/v) alginate was slightly higher than those prepared from 1.8 and 2.0% (w/v) alginate in cheese milk. Flavourzyme-alginate capsules prepared from 1.6, 1.8 and 2.0% (w/v) alginate retained over 70% of the initial enzyme activity under simulated cheese-press pressure. Concentration of alginate had no significant effect (p > 0.05) on the retention of encapsulated Flavourzyme when the capsules were pressed for 4h; however when the simulated cheese press duration increased to 8 and 16h the retention of encapsulated Flavourzyme was significantly higher (p [less than] 0.01) in capsules produced from 2.0% (w/v) alginate. Incorporation of encapsulated enzymes into the milk prior to rennetting resulted in an even distribution of capsules in the cheese matrix compared to aggregation of capsules, when added to milled curd prior to salting. All cheeses; control with no added enzymes and experimental cheeses with free and encapsulated Flavourzyme and/or Palatase showed higher levels of moisture and lower levels of fat compared to standard Cheddar cheese due to the variation in the manufacturing protocol. There was no significant difference (p > 0.05) in fat and final pH between control and experimental cheeses and there was no difference in the numbers of coliforms, E.coli, Salmonella, Listeria, coagulase positive staphylococci, Bacillus cereus, yeast and moulds in control or experimental cheeses. Increased and prolonged proteolysis was observed in cheeses with encapsulated Flavourzyme showing increased release of several peptides, also with the formation of new peptides absent in the control cheese with no added enzymes. Accumulation of high molecular weight/hydrophobic peptides was higher in cheeses with free Flavourzyme followed by cheeses with encapsulated Flavourzyme. Concentration of water-soluble peptides increased with the increase in the concentration of encapsulated Flavourzyme in the cheese. Concentration of water-insoluble peptides was higher in control cheese compared to cheeses with encapsulated Flavourzyme even after 180 days ripening. After 30 days of ripening, concentration of most free amino acids was about 3 times greater in cheeses with encapsulated Flavourzyme than in control and about 7 times higher after 90 days ripening. Concentration of total amino acids was consistently higher in cheeses with encapsulated Flavourzyme compared to control. Cheese grading scores for body, texture and appearance of all cheeses with encapsulated enzymes were lower than control and free enzyme treated cheeses during the entire grading period of about 100 days due to crumbly and pasty texture. Control and cheeses with added Flavourzyme received high overall score for flavour. Flavour score of cheese with encapsulated Flavourzyme at a concentration of 0.75 LAPU/g milk protein was higher than all cheeses around 50 days with better overall flavour score until about 94 days ripening with improved flavour and elimination of bitterness. However the flavour of enzyme treated cheeses deteriorated with time and the control cheese scored the highest for flavour. Though increased concentration of free fatty acids was detected in cheeses treated with encapsulated lipase; Palatase, these cheeses developed rancid, unpleasant, strong lipolytic flavours as early as 55 days ripening

Survival of co-encapsulated complementary probiotics and prebiotics in yoghurt

Complementary prebiotics for probiotic bacteria Lactobacillus acidophilus CSCC 2409 and Bifidobacterium infantis CSCC 1912 were selected by in vitro fermentation, and the efficacy of the prebiotics was tested by co-encapsulation with probiotic bacteria to improve survival in yoghurt. Raftilose for L. acidophilus CSCC 2409 (P0.05) were selected from in vitro fermentation studies. Yoghurts were prepared by incorporating probiotic cultures co-encapsulated with raftilose and stored for 6 weeks. Free cells of L. acidophilus CSCC 2409 and B. infantis 1912 showed a 3-log reduction in numbers in yoghurts at the end of the 6 week storage period. In contrast, L. acidophilus CSCC 2409 and B. infantis 1912 co-encapsulated with raftilose showed only 1-log reduction in numbers in yoghurts at the end of the 6 week storage period. This study demonstrates the potential protective effect of prebiotics such as raftilose in enhancing the survival of probiotic bacteria in yoghurt

Recent trends in accelerated cheese ripening using microencapsulated enzymes

Micro encapsulation is an inclusion technique for entrapping an active substance such as enzyme into a polymeric (gelled) matrix that may be coated by one or more semi-permeable polymers, by virtue of which the encapsulated compound becomes more stable than its free form

Impact of alginate-chitosan encapsulated flavourzyme on peptide and amino acid profiles in Cheddar cheese

The effect of alginate-chitosan encapsulated Flavourzyme on the water-soluble and water-insoluble peptides and amino acids released during Cheddar cheese ripening was investigated. Increased and prolonged proteolysis was observed in cheeses incorporated with encapsulated Flavourzyme. Rapid proteolysis and increased accumulation of hydrophobic or high molecular weight peptides was however observed in cheeses with free Flavourzyme compared to control (without Flavourzyme) or cheeses with encapsulated Flavourzyme. Concentration of watersoluble peptides increased with the concentration of encapsulated Flavourzyme in the cheese. Most free amino acids were about 3 times greater in cheese with encapsulated Flavourzyme compared to control cheese after 30 days ripening and about 7 times greater after 90 days ripening. Total amino acid content was highest in cheese with encapsulated Flavourzyme followed by free Flavourzyme treated cheese and lowest in control cheese. Flavourzyme encapsulated in slow release alginate-chitosan matrix can be a potential delivery system for flavour enhancement during cheese ripening

Circulating phospholipid profiling identifies portal contribution to NASH signature in obesity

International audienceBackground & AimsNonalcoholic steatohepatitis (NASH) is characterized by steatosis, lobular inflammation, hepatocyte ballooning with fibrosis in severe cases and high prevalence in obesity. We aimed at defining NASH signature in morbid obesity by mass spectrometry-based lipidomic analysis.MethodsSystemic blood before and 12 months post bariatric surgery along with portal blood and adipose tissue lipid efflux collected at the time of surgery from obese women were analyzed (9 structural classes, 150 species).ResultsIncreased concentrations of several Glycerophosphocholines (PC), Glycerophosphoethanolamines (PE), Glycerophosphoinositols (PI), Glycerophosphoglycerols (PG), Lyso-Glycerophosphocholines (LPC), and Ceramides (Cer) were detected in systemic circulation of NASH subjects. Weight loss post-surgery (12 months) improved the levels of liver enzymes, as well as several lipids, but most PG and Cer species remained elevated. Analysis of lipids from hepatic portal system at the time of surgery revealed limited lipid alterations compared to systemic circulation, but PG and PE classes were found significantly increased in NASH subjects. We evaluated the contribution of visceral adipose tissue to lipid alterations in portal circulation by measuring adipose tissue lipid efflux ex vivo, which demonstrated only minor alterations in NASH subjects. Interestingly, integration of clinical and lipidomic data (portal and systemic) led us to define a NASH signature in which lipids and clinical parameters are equal contributorsConclusionCirculatory (portal and systemic) phospholipid profiling and clinical data defines NASH signature in morbid obesity. We report weak contribution of visceral adipose tissue to NASH-related portal lipid alterations, suggesting possible contribution from other organs draining into hepatic portal system

A reference tissue atlas for the human kidney.

Kidney Precision Medicine Project (KPMP) is building a spatially specified human kidney tissue atlas in health and disease with single-cell resolution. Here, we describe the construction of an integrated reference map of cells, pathways, and genes using unaffected regions of nephrectomy tissues and undiseased human biopsies from 56 adult subjects. We use single-cell/nucleus transcriptomics, subsegmental laser microdissection transcriptomics and proteomics, near-single-cell proteomics, 3D and CODEX imaging, and spatial metabolomics to hierarchically identify genes, pathways, and cells. Integrated data from these different technologies coherently identify cell types/subtypes within different nephron segments and the interstitium. These profiles describe cell-level functional organization of the kidney following its physiological functions and link cell subtypes to genes, proteins, metabolites, and pathways. They further show that messenger RNA levels along the nephron are congruent with the subsegmental physiological activity. This reference atlas provides a framework for the classification of kidney disease when multiple molecular mechanisms underlie convergent clinical phenotypes