In order to determine the effect of lipid concentration on the properties of k-casein stabilized emulsions, butteroil was added to solutions that contained 0.3% k-casein to achieve milk fat concentrations of 3, 10, and 20%. These mixtures were adjusted to pH 6.5 and heated to 65°C. They were then homogenized at 20 and 100 MPa and particle size was measured; viscosity and yield stress were measured before and 30 minutes after the addition of chymosin. These experiments were repeated twice. Homogenization of the emulsions at 100 MPa produced smaller particles than homogenization at 20 MPa. Emulsions with 20% milk fat showed the largest particle size. Before treated with chymosin, these emulsions had the greatest viscosity and yield stress, however the differences with the other lipid concentrations were greater after chymosin treatment. A gel with yield stress less than 10 Pa occurred in emulsions with 3 or 10% milk fat. The emulsion with 20% milk fat after chymosin treatment provided the best possibility for the formation of a gel because it had the highest viscosity and yield stress. The effect of protein concentration on the properties of the emulsions was determined in emulsions that contained 20% milk fat and 0.5, 0.7, and 1.0%. These emulsions were prepared as previously described. Emulsions homogenized at 100 MPa had smaller particles than emulsions homogenized at 20 MPa. An increase in protein concentration caused the particle size to decrease. Emulsions homogenized at 100 MPa were more stable than emulsions homogenized at 20 MPa and the emulsion with 1.0% k-casein was the most stable emulsion. The protein load of k-casein stabilized emulsions ranged from 3 to 6 mg/m2. The viscosity and yield stress prior to chymosin treatment showed no properties of gelation. After treated with chymosin, these emulsions produced a weak gel with yield stress values that ranged from 14 to 16 Pa.

No description available.

Several studies have demonstrated that certain genetic variants of beta-casein are closely related to milk production, milk composition and technological properties of milk such as coagulation properties during cheese making, calcium precipitation and water binding properties. The objective of current study is to investigate the effect of the genetic variants on hydrolysis of beta-casein by chymosin and pepsin. On the basis of a preliminary analysis, a total of 50 milk samples which provided representatives of different available genetic variants of beta-casein were collected from different herds in Quebec. Casein was prepared from milk samples by the acid precipitation and the genetic variants of beta-casein were identified by both alkaline and acid urea-PAGE. An anion exchange chromatography was employed for the separation of beta-casein from whole casein. An initial hydrolysis of beta-casein of different phenotypes by chymosin and pepsin were achieved under the optimized hydrolytic conditions. Hydrolysates were periodically removed from the reaction mixture and they were analyzed by both RP-HPLC and SDS-PAGE in order to study the hydrolytic pattern of each beta-casein variant with the increasing hydrolytic time. (Abstract shortened by UMI.)

Caseins are present in milk in the form of large spherical complexes called micelles that are stabilized by kappa-casein found on the surface. This stabilizing effect is lost when milk clotting enzymes hydrolyze kappa-casein thereby initiating the coagulation process. The objective of this study was to analyze the effect of kappa-casein polymorphism upon hydrolysis by proteases. The A and B forms are the most common genetic variants of kappa-casein in Canadian Holstein dairy herds, representing three phenotypes AA, AB and BB. Whole casein from Holstein milk samples was fractionated into its four major components by ion-exchange chromatography using Express Ion Exchanger Q Anion Exchanger and Macro-Prep High Q Anion Exchange Support columns. The kappa-casein fraction was isolated, dialyzed, and assessed for its phenotype and purity by PAGE. The pure forms of the three different phenotypes of kappa-casein (AA, AB, BB) at a final concentration of 0.5% were then hydrolyzed by calf chymosin (1:500) and porcine pepsin (1:1000) at a pH of 5.8 at 37°C. Aliquots were collected at 0, 5, 15, 30, 60 and 90 min and the reaction stopped by using 24% NH4OH. The rate of hydrolysis of the three phenotypes was analyzed by comparing the disappearance of the substrate with time from the RP-HPLC chromatograms using Waters MAXIMA software. SDS-PAGE was used to calculate the approximate molecular weight of the hydrolytic products. Analysis of the hydrolysate profiles indicated that there was significant difference (P < 0.05) in the rate of hydrolysis between the phenotypes. Under the present conditions at a pH of 5.8, the AA phenotype showed a significantly slower rate of hydrolysis by both chymosin and pepsin, than the other two phenotypes. There was no significant difference in the rate of hydrolysis between the phenotypes AB and BB during chymosin hydrolysis. The BB phenotype is hydrolyzed more extensively and the AB phenotype is intermediate between the two variants in the p

No description available.

No description available.

Die Dicklegung der Milch, ausgehend von der hydrolytischen Spaltung des κ-Caseins, stellt den ersten essentiellen Schritt in der Käseherstellung dar. Dabei finden Gerinnungsenzyme verschiedener Herkunft Anwendung, deren neueste Varianten auf Grundlage des aktuellen Forschungsstandes umfassend charakterisiert werden. Verschiedene Kälberlabpräparate, mikrobielle Gerinnungsenzyme aus Rhizomucor miehei und mit Hilfe von gentechnisch modifizierten Mikroorganismen gewonnenes Chymosin (FPC) aus Rind und Kamel werden mittels HPLC und Elektrophorese hinsichtlich ihrer Zusammensetzung analysiert. Die neueste Generation mikrobieller Enzyme weist im Gegensatz zur herkömmlichen Variante keine Nebenkomponenten und damit einen höheren Aufreinigungsgrad auf. Die unspezifische proteolytische Aktivität wird durch fluorimetrische Quantifizierung der in 12 % TCA löslichen Stickstoffkomponenten bestimmt, die nach Inkubation rekonstituierter Magermilch bei 32 °C und pH 6,5 über 24 h mit den Enzymen freigesetzt werden. Mikrobielle Gerinnungsenzyme besitzen eine signifikant höhere unspezifische proteolytische Aktivität gegenüber chymosinbasierten Präparaten, deren Auswirkung sich bei Erhöhung der zugegebenen Enzymmenge besonders ausgeprägt darstellen. Oszillationsrheometrische Analysen lassen bei gleicher Enzymaktivität eine geringere Gelfestigkeit nach 40 min bei Einsatz von mikrobiellen Präparaten im Vergleich zu Kälberlab und bovinem FPC erkennen. Zusätzlich wird eine Abhängigkeit der Flockungszeit und der Gelfestigkeit vom eingesetzten Substrat beobachtet, die für Chymosin aus Kamel besonders stark ausgeprägt ist. Die Substratspezifität dieses Enzyms ist weder mit der des bovinen Chymosins noch mit der der mikrobiellen Gerinnungsenzyme identisch. Im Rahmen von Käsungsversuchen im Labor-, Pilot- und Industriemaßstab wird eine signifikant höhere Käseausbeute (0,50 - 1,19 %) bei Verwendung vom traditionellem Kälberlab im Vergleich zur neuesten Generation der kommerziellen mikrobiellen Substitute ermittelt. Im Verlaufe der Reifung von Schnittkäse wird durch mikrobielles Gerinnungsenzym gegenüber Kälberlab eine signifikant höhere Menge an Nichtproteinstickstoff freigesetzt sowie ein unterschiedliches Profil an Proteinabbauprodukten gebildet. Die höhere proteolytische Aktivität resultiert in einer signifikant stärker ausgeprägten Bitterkeit der mit mikrobiellem Gerinnungsenzym hergestellten Käse nach 12 Wochen Reifungszeit. / Clotting of milk caused by hydrolytic cleavage of κ-casein is the first important step in cheese milk processing. This cleavage is caused by clotting enzymes of different origin, which are comprehensively characterized by considering results of latest investigations. The composition of selected calf rennets, microbial coagulants derived from Rhizomucor miehei and genetically engineered chymosin (FPC) derived from cow and camel is analyzed by HPLC and electrophoresis. In contrast to conventional products, the latest generation of microbial coagulants does not show minor components in a detectable amount because of a sufficient purification. The unspecific proteolytic activity is determined by fluorimetric quantification of 12 % tricloric-acid-soluble nitrogen, which is released by the enzymes from reconstituted skim milk, pH 6.5, after incubation at 32 °C for 24 h. Microbial coagulants show a significantly higher unspecific proteolysis as compared to chymosin-based clotting enzymes, especially when the enzymes are added in amount higher than used during cheese-making. Small amplitude oscillation rheometry analysis showed a lower gel firmness after 40 min of gelling when microbial coagulants were applied instead of calf rennet or FPC. Furthermore, flocculation time, gel formation time and gel firmness additionally depends on the test substrate, and this dependency is exceptionally pronounced when camel chymosin was used. The substrate specificity of this enzyme is neither identical to that of bovine chymosin nor to that of microbial coagulants. Cheese making experiments in laboratory-, pilot- and commercial-scale revealed a significantly higher cheese yield (0.50 - 1.19 %) when using calf rennet instead of microbial coagulant of the latest generation. During ripening of semi-hard cheese a higher amount of non-protein-nitrogen was released and a different electrophoretic casein degradation profile was generated when using microbial enzymes. Enhanced proteolysis is responsible for a significantly higher pronounced bitterness of microbial produced cheese after 12 weeks of maturation.

Die Dicklegung der Milch, ausgehend von der hydrolytischen Spaltung des κ-Caseins, stellt den ersten essentiellen Schritt in der Käseherstellung dar. Dabei finden Gerinnungsenzyme verschiedener Herkunft Anwendung, deren neueste Varianten auf Grundlage des aktuellen Forschungsstandes umfassend charakterisiert werden. Verschiedene Kälberlabpräparate, mikrobielle Gerinnungsenzyme aus Rhizomucor miehei und mit Hilfe von gentechnisch modifizierten Mikroorganismen gewonnenes Chymosin (FPC) aus Rind und Kamel werden mittels HPLC und Elektrophorese hinsichtlich ihrer Zusammensetzung analysiert. Die neueste Generation mikrobieller Enzyme weist im Gegensatz zur herkömmlichen Variante keine Nebenkomponenten und damit einen höheren Aufreinigungsgrad auf. Die unspezifische proteolytische Aktivität wird durch fluorimetrische Quantifizierung der in 12 % TCA löslichen Stickstoffkomponenten bestimmt, die nach Inkubation rekonstituierter Magermilch bei 32 °C und pH 6,5 über 24 h mit den Enzymen freigesetzt werden. Mikrobielle Gerinnungsenzyme besitzen eine signifikant höhere unspezifische proteolytische Aktivität gegenüber chymosinbasierten Präparaten, deren Auswirkung sich bei Erhöhung der zugegebenen Enzymmenge besonders ausgeprägt darstellen. Oszillationsrheometrische Analysen lassen bei gleicher Enzymaktivität eine geringere Gelfestigkeit nach 40 min bei Einsatz von mikrobiellen Präparaten im Vergleich zu Kälberlab und bovinem FPC erkennen. Zusätzlich wird eine Abhängigkeit der Flockungszeit und der Gelfestigkeit vom eingesetzten Substrat beobachtet, die für Chymosin aus Kamel besonders stark ausgeprägt ist. Die Substratspezifität dieses Enzyms ist weder mit der des bovinen Chymosins noch mit der der mikrobiellen Gerinnungsenzyme identisch. Im Rahmen von Käsungsversuchen im Labor-, Pilot- und Industriemaßstab wird eine signifikant höhere Käseausbeute (0,50 - 1,19 %) bei Verwendung vom traditionellem Kälberlab im Vergleich zur neuesten Generation der kommerziellen mikrobiellen Substitute ermittelt. Im Verlaufe der Reifung von Schnittkäse wird durch mikrobielles Gerinnungsenzym gegenüber Kälberlab eine signifikant höhere Menge an Nichtproteinstickstoff freigesetzt sowie ein unterschiedliches Profil an Proteinabbauprodukten gebildet. Die höhere proteolytische Aktivität resultiert in einer signifikant stärker ausgeprägten Bitterkeit der mit mikrobiellem Gerinnungsenzym hergestellten Käse nach 12 Wochen Reifungszeit. / Clotting of milk caused by hydrolytic cleavage of κ-casein is the first important step in cheese milk processing. This cleavage is caused by clotting enzymes of different origin, which are comprehensively characterized by considering results of latest investigations. The composition of selected calf rennets, microbial coagulants derived from Rhizomucor miehei and genetically engineered chymosin (FPC) derived from cow and camel is analyzed by HPLC and electrophoresis. In contrast to conventional products, the latest generation of microbial coagulants does not show minor components in a detectable amount because of a sufficient purification. The unspecific proteolytic activity is determined by fluorimetric quantification of 12 % tricloric-acid-soluble nitrogen, which is released by the enzymes from reconstituted skim milk, pH 6.5, after incubation at 32 °C for 24 h. Microbial coagulants show a significantly higher unspecific proteolysis as compared to chymosin-based clotting enzymes, especially when the enzymes are added in amount higher than used during cheese-making. Small amplitude oscillation rheometry analysis showed a lower gel firmness after 40 min of gelling when microbial coagulants were applied instead of calf rennet or FPC. Furthermore, flocculation time, gel formation time and gel firmness additionally depends on the test substrate, and this dependency is exceptionally pronounced when camel chymosin was used. The substrate specificity of this enzyme is neither identical to that of bovine chymosin nor to that of microbial coagulants. Cheese making experiments in laboratory-, pilot- and commercial-scale revealed a significantly higher cheese yield (0.50 - 1.19 %) when using calf rennet instead of microbial coagulant of the latest generation. During ripening of semi-hard cheese a higher amount of non-protein-nitrogen was released and a different electrophoretic casein degradation profile was generated when using microbial enzymes. Enhanced proteolysis is responsible for a significantly higher pronounced bitterness of microbial produced cheese after 12 weeks of maturation.

Diss. (sammanfattning) Uppsala : Sveriges lantbruksuniv., 2008. / Härtill 5 uppsatser.

<p><strong>Apstrakt je obrađen tehnologijama za optičko prepoznavanje teksta (OCR).</strong></p><p>Imajući u vidu naučni i praktičan značaj i nedostatak podataka o aplikaciji ultrafiltracije (UF), alternativnih koaguli&scaron;ućih enzima, posebno genetskog himozina, kao i enzima za akceleraciju zrenja u tehnologiji tvrdih sireva, za predmetna istraživanja je odabran Kačkavalj, popularni sir mediteranskog porekla.<br />U okviru istraživanja je u industrijskim uslovima izvr&scaron;ena proizvodnja KačkavaIja iz retentata dobijenog nakon ultrafiltracije mleka po LCR (Low Concentrated Retentate) postupku (UF Kačkavalj). Ispitan je uticaj koagulanasa mikrobiolo&scaron;kog porekla (M. miehei proteaza, Rennilase) i genetski modifikovanog himozina (Maxiren) na kvalitet finalnog sira i upoređen sa klasičnim analozima (kontrolni Kačkavalj). Istražena je mogućnost skraćenja perioda zrenja obe varijante Kačkavalja (klasični i UF) dodatkom &bdquo;koktela&quot; enzima za ubrzanje zrenja proteolitičkog (Accelase) i lipolitičkog (Paletase) dejstva.<br />Na osnovu rezultata dobijenih ispitivanjem fizičko-hemijskog sastava, mikro-strukture i senzornog kvaliteta varijanti konvencionalnog Kačkavalja i UF analoga tokom jednogodi&scaron;njeg zrenja, utvrđeno je da se dejstvo rekombinovanog himozina (Maxiren-a) i standardnog himozina signifikantno ne razlikuje, potvrđujući dosada&scaron;nja saznanja da je genetski modifikovani himozin potpuno kompetentan proteolitički agens za koagulaciju mleka u procesu proizvodnje sira. Kod konvencionalnog i UF Kačkavalja proizvedenog primenom mikrobiolo&scaron;ke M. miehei proteaze (Rennilase) ustanovljena je najniža koncentracija mlečne masti u suvoj materiji tokom celokupnog ispitivanog perioda, &scaron;to se sa ekonomskog aspekta negativno odražava na kvalitet sira. Sve varijante UF Kačkavalja se tokom zrenja razlikuju od klasičnih analoga po hemijskom sastavu (sadržaj mlečne masti, ukupnog N, NPN, RN itd.), profilu proteolitičke degradacije as- i p-kazeina, modelu distribucije identifikovanih is- parljivih komponenata arome, reolo&scaron;kim i senzornim karakteristikama i mikrostrukturi usled<br />usporenog intenziteta promena proteinske i lipidne faze UF sireva tokom zrenja. Efekat ubrzanja zrenja Kačkavalja tretiranog &bdquo;koktelom&quot; enzima lipolitičkog i proteolitičkog dejstva je izraženiji kod konvencionalnog nego UF uzorka.</p> / <p><strong>Abstract was processed by technology for Optical character recognition (OCR).</strong></p><p>The scientific and practical significance and the lack of literature data on the<br />application of ultrafiltration (UF), as well as the alternative coagulating enzymes, particularly genetic chymosin, and enzymes for accelerated ripening in hard cheese technology were essential reasons for choosing this field for research. The object of presented investigations was Kashkaval, the popular mediterranean hard variety cheese.<br />In the scope of these investigations, Kashkaval was industrially produced using retentate obtained by UF of milk according to LCR (Low Concentrated Retentate) procedure (UF Kashkaval). Effects of clotting enzymes of microbiological origine (M. miehei protease, Rennilase) and of genetically modified chymosin (Maxiren) on the quality of final cheese product was also investigated and compared to conventional analogues (control Kashkaval). The possibility for reduced ripening period of both Kashkaval types (traditional and UF) was investigated by the addition of &bdquo;coctail&quot; of enzymes which accelerate the cheese ripening, involving the proteolytic (Accelase) and lypolytic (Palatase) activity.<br />According to the results obtained by the investigations of physico-chemical composition, microstructure and sensory quality of conventional Kashkaval and UF analogues in the course of one-year ripening period, it was possible to derive the conclusion that the effects of recombined chymosin (Maxiren) and the standard one are insignificantly different, confirming thus the former knowledge that the genetically modified chymosin is an adequate proteolytic agent for milk coagulation in cheese production. The lowest level of milk fat in dry matter over the whole investigated period was established both for conventional and UF Kashkaval samples produced by the application of M. miehei protease (Rennilase), whereas from the economic standpoint this indicated negative effects on cheese quality. All UF Kashkaval varieties differ during the period of ripening from conventional analogues concerning chemical composition (content of milk fat, total N, NPN, RN etc.), profile of proteolytic degradation of as- and p-casein, distribution of identified volatile aroma<br />components, rheological and sensory characteristics as well as the microstructure due to the slower intensity of changes in protein and lipid phases of UF cheese samples during ripening. The &bdquo;cocktail&quot; of proteolytic and lipolytic enzymes showed more pronounced effects on the Kashkaval ripening in the case of conventionally produced cheese sample.</p>