Made available in DSpace on 2014-06-11T19:32:54Z (GMT). No. of bitstreams: 0 Previous issue date: 2007-04-05Bitstream added on 2014-06-13T20:44:16Z : No. of bitstreams: 1 silveira_gg_dr_rcla.pdf: 641894 bytes, checksum: 52f7995780eb514330fda7bbbcb68b69 (MD5) / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) / Neste trabalho foi estudada a produção de renina microbiana pelo zigomiceto Mucor miehei utilizando Fermentação em Estado Sólido (FES). Os substratos utilizados foram farelos de trigo, laranja, aveia e arroz, bagaço de cana e casca de café. Foi estudada também a combinação de várias quantidades de dois substratos diferentes já mencionados. Dos substratos utilizados, não houve crescimento evidenciado do fungo quando foi utilizado, casca de café, bagaço de cana e farelo de laranja. Caseína é um importante fator na potencialização da biosíntese de renina tanto em fermentação submersa quanto em Fermentação em Estado Sólido. Nos experimentos com farelos de trigo e aveia as maiores atividades enzimáticas foram observadas entre 48 e 72 horas de cultivo, umidade acima de 79% e o aumento da quantidade de farelo de trigo na mistura foi favorável no que diz respeito ao aumento desta atividade. Nos experimentos com farelos de trigo e arroz sem suplementação de caseína, todas as combinações de meio de cultivo foram estatisticamente significativas sobre a atividade enzimática. Umidades acima de 98 % parecem inibir o crescimento do fungo e, consequentemente, a síntese enzimática que aumentou após 48 horas de cultivo. Quando houve a suplementação de caseína nos experimentos com farelos de trigo e arroz, houve um sensível incremento nas atividades enzimáticas. Os meios 2, 3 e 4 não forneceram influência estatisticamente importantes segundo o teste de Tukey e as menores atividades foram observadas no meio tipo 1. Umidades abaixo de 80 % e acima de 100 % forneceram baixas atividades enzimáticas. Assim como nos experimentos anteriores, as maiores atividades foram observadas após 48 horas de cultivo. Analisando-se todos os experimentos, verifica-se que as maiores atividades enzimáticas foram superiores quando comparadas aos relatos de outros pesquisadores. / The microbial rennin production by zigomiceto Mucor miehei produced on Solid State Fermentation (SSF) using wheat bran, rice bran, oat bran, sugarcane bagasse and coffee husk was studied in this work. The combination of two different substrates was also studied. No growth of microorganism was observed when coffee husk, sugar cane bagasse and orange pulp were used. Casein has an important factor in the rennin biosynthesis induction in Submerged Fermentation and Solid State Fermentation. In the experiments with wheat bran and oat bran the maximum enzyme activities had been observed between 48 and 72 hours. Moisture above of 79% and the wheat bran increase in the mixture were favorable for increase enzyme activity. Moisture above 98 % can inhibit the microorganism growth and the enzyme synthesis increased after 48 hours. When casein was added in wheat bran and rice bran medium, a rise in the enzymatic activities was observed. Low enzyme activities were observed in moisture between 80 % and 100 %. The highest activity was observed after 48 hours. The enzyme activity in this study was upper when compared with other researchers.

Rennet extract is an important commercial enzyme preparation which is indispensable to the cheese making industry, The term "rennet" is used to denote the enzyme system extracted from the fourth stomach (abomasum) of a suckling calf. Most of the enzymatic material in the extract is rennin. It also contains a small amount of pepsin and some other proteolytic enzymes. The amount of pepsin in rennet extract appears to depend upon the age of the calves at the time of slaughter. It has been suggested that some proteolytic enzymes from higher plants might be useful in cheese manufacture. Krishnaswamy et al. reported that Cheddar cheese made with ficin quickly developed a bitter flavor which decreased in intensity during curing. Cheddar cheese made with an extract from the flower petals of Cynara cardunculus developed extreme bitterness and a pasty body during 30 days curing at 10 C. An extract from the fruit of Withania coagulans was used by Kothavalla and Khubchandani as a coagulating agent in the manufacture of Surati and Cheddar cheese. No unusual flavors were reported, but they experienced rather high fat losses.

A procedure was developed for measuring residual rennin activity in curd extracts and whey. A sensitive substrate at pH 5.8 was prepared by mixing 6 grams of low-heat nonfat dry milk in 500 milliliters buffer containing 0.05 molar cacodylic acid, 0.02 molar calcium chloride, and 0.012 molar triethanolamine, and storing it at 2 degrees centigrade for 18 hours. Two milliliters of whey or curd extract were inoculated into 25 milliliters of substrate at 30 degrees centigrade. The coagulation time was measured and compared to that induced by 2 milliliters of a known rennin concentration added at the same time to identical substrate. Recovery of activity was determined by adjusting milk samples to pH 5.20 and coagulating them with a known concentration of rennin. The clot was broken by agitation and the curd separated from the whey by centrifugation. Activities accounted for in the curd and whey amounted to 91±1.6 per cent the activity added to the milk. Maximum release of activity from curd was achieved by diluting the curd 1:15 with water and adjusting the pH to 6.80. Inclusion of curd particles with the inoculum decreased the sensitivity of the substrate.

Thesis--Copenhagen University. / Summary in Danish. Includes index. Bibliography: p. 139-162.

Resumo: Neste trabalho foi estudada a produção de renina microbiana pelo zigomiceto Mucor miehei utilizando Fermentação em Estado Sólido (FES). Os substratos utilizados foram farelos de trigo, laranja, aveia e arroz, bagaço de cana e casca de café. Foi estudada também a combinação de várias quantidades de dois substratos diferentes já mencionados. Dos substratos utilizados, não houve crescimento evidenciado do fungo quando foi utilizado, casca de café, bagaço de cana e farelo de laranja. Caseína é um importante fator na potencialização da biosíntese de renina tanto em fermentação submersa quanto em Fermentação em Estado Sólido. Nos experimentos com farelos de trigo e aveia as maiores atividades enzimáticas foram observadas entre 48 e 72 horas de cultivo, umidade acima de 79% e o aumento da quantidade de farelo de trigo na mistura foi favorável no que diz respeito ao aumento desta atividade. Nos experimentos com farelos de trigo e arroz sem suplementação de caseína, todas as combinações de meio de cultivo foram estatisticamente significativas sobre a atividade enzimática. Umidades acima de 98 % parecem inibir o crescimento do fungo e, consequentemente, a síntese enzimática que aumentou após 48 horas de cultivo. Quando houve a suplementação de caseína nos experimentos com farelos de trigo e arroz, houve um sensível incremento nas atividades enzimáticas. Os meios 2, 3 e 4 não forneceram influência estatisticamente importantes segundo o teste de Tukey e as menores atividades foram observadas no meio tipo 1. Umidades abaixo de 80 % e acima de 100 % forneceram baixas atividades enzimáticas. Assim como nos experimentos anteriores, as maiores atividades foram observadas após 48 horas de cultivo. Analisando-se todos os experimentos, verifica-se que as maiores atividades enzimáticas foram superiores quando comparadas aos relatos de outros pesquisadores. / Abstract: The microbial rennin production by zigomiceto Mucor miehei produced on Solid State Fermentation (SSF) using wheat bran, rice bran, oat bran, sugarcane bagasse and coffee husk was studied in this work. The combination of two different substrates was also studied. No growth of microorganism was observed when coffee husk, sugar cane bagasse and orange pulp were used. Casein has an important factor in the rennin biosynthesis induction in Submerged Fermentation and Solid State Fermentation. In the experiments with wheat bran and oat bran the maximum enzyme activities had been observed between 48 and 72 hours. Moisture above of 79% and the wheat bran increase in the mixture were favorable for increase enzyme activity. Moisture above 98 % can inhibit the microorganism growth and the enzyme synthesis increased after 48 hours. When casein was added in wheat bran and rice bran medium, a rise in the enzymatic activities was observed. Low enzyme activities were observed in moisture between 80 % and 100 %. The highest activity was observed after 48 hours. The enzyme activity in this study was upper when compared with other researchers. / Orientador: Jonas Contiero / Coorientador: Antonio Carlos Simões Pião / Banca: Dejanira de Franceschi de Angelis / Banca: Eloízio Julio Ribeiro / Banca: Jorge José Correa Lopes / Banca: José Sílvio Govone / Doutor

A procedure was developed for the extraction of rennet from fresh frozen vells. Frozen vells were minced in a Model VCM-25 Hobart Vertical Cutter/Mixer. Dry sodium chloride was added until the salt concentration in the moisture of the tissue was 10%. Salted tissue was placed in galvanized steel cells and heated in a water bath to 49, 54, 57 and 60 C for 0, 15, 30, 45 and 60 minutes. The heated tissue was dried to about 4% moisture at ambient temperature with the aid of an electric fan. The dried salted tissue was extracted with a sodium chloride solution adjusted to give a salt concentration of 10% in the extraction solution. Salted vell tissue heated to 49 C for 60 minutes and to 54 C for 30 minutes prior to drying produced 82 and 55%, respectively, more recoverable enzyme activity than unheated salted tissue. However, tissue which was salted, heated and not dried prior to extraction produced less recoverable activity than unheated samples. Unsalted vell tissue which was heated to 49, 54, 57 and 60 C for 15, 30, 45 and 60 minutes and extracted without being dried yielded lower recoverable activity than unheated samples. Heating unsalted vell tissue to 54, 57 and 60 C for 15, 30, 45 and 60 minutes prior to drying did not increase recoverable enzyme activity over unheated samples. Unsalted tissue heated to 49 C produced a slightly higher recovery than unheated samples.

Residual rennin in cheese whey and curd was measured by using a special sensitive substrate. The substrate was made by reconstituting 6 g NDM in 500 ml of buffer containing 0.2 M CaCl2, 0.5 M cacodylic acid and 0.2 M triethanolamine at pH 5.7. Cheese curd was blended into a 1:7 slurry (1 part curd, 7 parts water), and 1.67% sodium chloride was added to the why and slurry to liberate residual rennin from casein. The residual rennin in cheese whey and slurry were determined simultaneously with an identical sample containing known rennin activity. Samples with known activity were prepared by destroying the residual rennin in unknown samples after which a known amount of rennin was added back to a standard. The examine the effectiveness of this method for measuring rennin activity in whey or slurry, a recovery test was developed to measure rennin activities in the whey and curd made by centrifuging rennet-coagulated milk. The average total recovery from 15 replications was 101.6 ± 2.4%. It was found that pH was a main factor affecting rennin distribution between whey and curd. The amount in the curd increased with decreasing pH at setting. Adding 0.02% CaCl2 to milk was of little effect. The results showed that 68.1 6 ± 6.6% of the residual rennin was found in Cheddar cheese whey after dipping and 17.2 6 ± 2.6% of the residual rennin was in the curd after milling.

Thesis--Copenhagen University. / Summary in Danish. Includes index. Bibliography: p. 139-162.