Ciências Exatas e Tecnológicas
URI permanente desta comunidadehttps://locus.ufv.br/handle/123456789/4
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Item Ultra high temperature milk: microbiological quality, age gelation and a proposal of an alternative method for microbial detection(Universidade Federal de Viçosa, 2024-02-27) Moreira, Isabella Maria Fernandes Botelho; Carvalho, Antônio Fernandes de; http://lattes.cnpq.br/5311708809706410The UHT (Ultra High Temperature) milk is a food that has stood out in the market due to its ease of transport, storage conditions and long shelf life. Due to this, it is becoming increasingly important to study this product in order to guarantee the quality and food safety of the consumer. The objective of this study was to evaluate three different lines within the “UHT milk universe”: the microbiological quality of the product, the action of stabilizing salts on the gelation process caused by bacterial proteases and an alternative and faster methodology for microbial detection. Microbiological quality was determined through the evaluation of 184 samples of Brazilian UHT milk for the presence of vegetative cells of aerobic mesophilic bacteria and spores. The 335 isolates obtained were purified, phenotypically characterized and subjected to molecular analyzes in order to determine the biodiversity between them. By constructing a phylogenetic tree, it was possible to identify bacteria belonging to the genus Bacillus sp. that in addition to being capable of forming spores, some strains in the group also secrete proteolytic and lipolytic enzymes that compromise the quality of the product throughout its shelf life. Based on this, the second project aimed to evaluate the action of phosphate and citrate salts on the gelation process in UHT milk samples intentionally inoculated with strains of Pseudomonas fluorescens 07A, a strain that produces the AprX protease, for a period of 18 h at 20 °C. In this project, five stabilizing salts (polyphosphate (T1), pyrophosphate (T2), citrate (T3), tripolyphosphate (T4) and monophosphate (T5)) were evaluated for 30 days in three repetitions in samples of UHT milk produced through reconstitution of skimmed milk powder. Through physical-chemical analyzes (pH, thermal stability in the oil bath, azocasein assay, SDS-page, protein content) and visual evaluation of the gel formed at the bottom of the sample bottles, it was possible to determine that the treatment added with tripolyphosphate (T4) was that showed greater efficiency in reducing the speed gelation of samples. Finally, the last project came from the demand for less laborious and faster methods for detecting microbial contamination in food. The BD BACTEC™ FX is currently used for the rapid detection of microbial growth in blood samples. The project was divided into two stages in which, in the first, samples of UHT milk intentionally inoculated with strains of Bacillus spizizenii ATCC19659 were evaluated, and in the second, industrial samples of UHT milk were evaluated. Through the results obtained, it was verified that the equipment was very promising due to its ease of use, shorter contamination detection time and greater sensitivity when compared to conventional methodologies. The results obtained in all three projects were very satisfactory, but they are just the beginning of a range of possibilities for advances and discoveries that permeate the main object of the study, UHT milk. Keywords: UHT milk; Quality; Age Gelation; Stabilizing Salts; Alternative Analysis Methods.Item Microbiological quality of brazilian UHT milk and genetic biodiversity of spore-forming bacteria(Universidade Federal de Viçosa, 2019-02-21) Moreira, Isabella Maria Fernandes Botelho; Carvalho, Antônio Fernandes de; http://lattes.cnpq.br/5311708809706410Milk is considered a food rich in nutrients, which makes it the ideal environment for the development of a series of microorganisms. Among the milk contaminants microorganisms, which are able to form heat-resistant spores are very important, especially for dairy companies that use UHT technology. The genus Bacillus is the most associated with milk quality, and between the species of importance in the dairy industry, we can highlight Bacillus sporothermodurans. This bacterium is an aerobic mesophyll able to produce spores resistant to UHT treatment, and throughout the storage can germinate and multiply until counts close to 105 CFU∙mL-1. The objective of this study was to determine the microbiological quality of UHT milk commercialized in Brazil with a focus on the biodiversity of sporulated thermoresistant microorganisms. This work began with the evaluation of samples of UHT milk donated by seven different companies (A, B, C, D, E, F and G), of three regions of the country (South, Southeast and Midwest) over six months. For that the steps of incubation and total plate count were performed in accordance with the Ordinance no 370 of 1997 of Ministério da Agricultura, Pecuária e Abastecimento. According to this Ordinance, which establishes that the count of aerobic mesophylls can not exceed the limit of 100 CFU∙mL-1, 50,5% of the samples evaluated were out of standards. After the seven days of incubation, two heat treatments (T1: 80 oC / 10 min and T2: 100 oC / 30 min) were applied to the inactivation of the vegetative cells and evaluation of the presence of sporulates. About five colonies were isolated from each sample and submitted to phenotypic characterization to identify the presence of B. sporothermodurans. For this, the analyzes were based on Normative Instruction n° 62 of 2003 of Ministério da Agricultura, Pecuária e Abastecimento in which each purified isolate was submitted to a series of biochemical tests (Gram staining, catalase, oxidase, anaerobic growth, glucose fermentation, esculin hydrolysis, nitrate reduction and urease test). According to the test results, none of the isolates obtained was considered B. sporothermodurans. Subsequently, the Rep-PCR analysis was carried out, which allowed grouping the isolates into clusters. After choosing, randomly, the representatives of each group, considering a similarity of 90%, these were sent to the 16S rDNA sequencing and it can be observed that the genus Bacillus was found in abundance corresponding to 100% of the isolates. Within the genus Bacillus six species were identified: Bacillus amyloliquefaciens, Bacillus proteolyticus, Bacillus cereus, Bacillus subtilis, Bacillus licheniformis and Bacillus velezensis. The identification of the presence of these sporulated bacteria in UHT milk samples indicates the importance of quality maintenance of raw material, the implementation of self-control programs, and the awareness of all links in the production chain in order to obtain a quality product.Item Microbiological quality of brazilian UHT Milk and genetic biodiversity of spore-forming bacteria(Universidade Federal de Viçosa, 2019-02-21) Moreira, Isabella Maria Fernandes Botelho; Carvalho, Antônio Fernandes de; http://lattes.cnpq.br/5311708809706410Milk is considered a food rich in nutrients, which makes it the ideal environment for the development of a series of microorganisms. Among the milk contaminants microorganisms, which are able to form heat-resistant spores are very important, especially for dairy companies that use UHT technology. The genus Bacillus is the most associated with milk quality, and between the species of importance in the dairy industry, we can highlight Bacillus sporothermodurans. This bacterium is an aerobic mesophyll able to produce spores resistant to UHT treatment, and throughout the storage can germinate and multiply until counts close to 105 CFU∙mL-1. The objective of this study was to determine the microbiological quality of UHT milk commercialized in Brazil with a focus on the biodiversity of sporulated thermoresistant microorganisms. This work began with the evaluation of samples of UHT milk donated by seven different companies (A, B, C, D, E, F and G), of three regions of the country (South, Southeast and Midwest) over six months. For that the steps of incubation and total plate count were performed in accordance with the Ordinance no 370 of 1997 of Ministério da Agricultura, Pecuária e Abastecimento. According to this Ordinance, which establishes that the count of aerobic mesophylls can not exceed the limit of 100 CFU∙mL-1, 50,5% of the samples evaluated were out of standards. After the seven days of incubation, two heat treatments (T1: 80 oC / 10 min and T2: 100 oC / 30 min) were applied to the inactivation of the vegetative cells and evaluation of the presence of sporulates. About five colonies were isolated from each sample and submitted to phenotypic characterization to identify the presence of B. sporothermodurans. For this, the analyzes were based on Normative Instruction n° 62 of 2003 of Ministério da Agricultura, Pecuária e Abastecimento in which each purified isolate was submitted to a series of biochemical tests (Gram staining, catalase, oxidase, anaerobic growth, glucose fermentation, esculin hydrolysis, nitrate reduction and urease test). According to the test results, none of the isolates obtained was considered B. sporothermodurans. Subsequently, the Rep-PCR analysis was carried out, which allowed grouping the isolates into clusters. After choosing, randomly, the representatives of each group, considering a similarity of 90%, these were sent to the 16S rDNA sequencing and it can be observed that the genus Bacillus was found in abundance corresponding to 100% of the isolates. Within the genus Bacillus six species were identified: Bacillus amyloliquefaciens, Bacillus proteolyticus, Bacillus cereus, Bacillus subtilis, Bacillus licheniformis and Bacillus velezensis. The identification of the presence of these sporulated bacteria in UHT milk samples indicates the importance of quality maintenance of raw material, the implementation of self-control programs, and the awareness of all links in the production chain in order to obtain a quality product.