Statistical optimisation of cell growth and carotenoid production by Rhodotorula mucilaginosa

Sequential statistical methods were used to maximise carotenoid production by a strain of Rhodotorula mucilaginosa, isolated from the Brazilian ecosystem. Initially, a factorial 2(5-1) experimental design was used, and the variables were pH and the levels of glucose, yeast extract, MgSO4.7H2O and KH2PO4. The nitrogen source (yeast extract) was the most important variable in enhancing carotenoid production; MgSO4.7H2O and KH2PO4 had a negative influence. The initial pH had no significant effect on carotenoid and cell productions. We further investigated the effects of glucose and yeast extract effects, using a second-order central composite design (CCD) to optimise carotenoid production, which was adequately approximated with a full quadratic equation obtained from a two-factor-2-level design. The analysis of quadratic surfaces showed that after 5 days of cultivation at 25ºC, the maximum carotenoid concentration (745 µg l-1) was obtained with 15 g l-1 of yeast extract and 20 g l-1 of glucose. The maximum carotenoid production (152 µg g-1) was obtained with 5 g l-1 yeast extract and 10 g l-1 glucose. Carotenoid formation was more sensitive to changes in yeast extract concentration than to changes in glucose concentration. Maximum cell production was achieved with 15-17 g l-1 of yeast extract and 15-20 g l-1 of glucose.

Statistical optimisation of cell growth and carotenoid production by Rhodotorula mucilaginosa

Sequential statistical methods were used to maximise carotenoid production by a strain of Rhodotorula mucilaginosa, isolated from the Brazilian ecosystem. Initially, a factorial 2(5-1) experimental design was used, and the variables were pH and the levels of glucose, yeast extract, MgSO4.7H2O and KH2PO4. The nitrogen source (yeast extract) was the most important variable in enhancing carotenoid production; MgSO4.7H2O and KH2PO4 had a negative influence. The initial pH had no significant effect on carotenoid and cell productions. We further investigated the effects of glucose and yeast extract effects, using a second-order central composite design (CCD) to optimise carotenoid production, which was adequately approximated with a full quadratic equation obtained from a two-factor-2-level design. The analysis of quadratic surfaces showed that after 5 days of cultivation at 25ºC, the maximum carotenoid concentration (745 µg l-1) was obtained with 15 g l-1 of yeast extract and 20 g l-1 of glucose. The maximum carotenoid production (152 µg g-1) was obtained with 5 g l-1 yeast extract and 10 g l-1 glucose. Carotenoid formation was more sensitive to changes in yeast extract concentration than to changes in glucose concentration. Maximum cell production was achieved with 15-17 g l-1 of yeast extract and 15-20 g l-1 of glucose.

Selection and characterization of carotenoid-producing yeasts from Campinas region, Brazil

The objective of the present study was to select and identify yeasts from Brazil capable of producing carotenoids. Pigmented yeasts were isolated from soil, leaves, fruits, flowers and a processed product. The samples were incubated at 30ºC in Erlenmeyer flasks, containing YM broth. After 48 hours, they were inoculated in Petri dishes with YM agar, and incubated at 30ºC during 120 hours. The yeast colonies, which presented yellow to red coloration, were transferred to culture tubes containing YM agar, and incubated at 30ºC for 72 hours. Out of 242 samples, only five had yellow to red color at high intensity. These highly pigmented yeasts were re-isolated in Petri dishes with YM agar and then transferred to tubes with GPYM agar. Identification through morphological and reproduction characteristics, along with physiological and biochemical tests, classified four strains as R. mucilaginosa and one strain as R. graminis. The main carotenoids extracted from them were identified through HPLC analysis as beta-carotene and torulene. The strains showed potential as promising microorganisms for the commercial production of carotenoids.

Este trabalho teve como objetivo selecionar e identificar leveduras encontradas no Brasil capazes de produzir carotenóides. As leveduras pigmentadas foram isoladas de amostras de solos, folhas, frutos, flores e um alimento processado. As amostras foram colocadas em frascos de erlenmeyer, contendo meio de Extrato de Malte e Levedura (YM), e incubadas a 30ºC. Após 48 horas, as amostras foram inoculadas em placas de petri contendo meio YM ágar e incubadas a 30ºC por 120 horas. As colônias, que apresentaram coloração entre amarelo e vermelho, foram transferidas para os tubos de culturas, contendo meio YM ágar e incubadas a 30ºC por 72 horas. Das 242 amostras, somente cinco delas apresentaram coloração intensa entre amarelo e vermelho. Estas colônias de leveduras foram reisoladas, em placas de petri contendo YM ágar e, posteriormente, transferidas para tubos de ensaios contendo GPYM ágar. A identificação das leveduras, baseada nas características morfológicas, de reprodução, além dos testes fisiológicos e bioquímicos, classificou quatro linhagens como Rhodotorula mucilaginosa e uma como Rhodotorula graminis. Os principais pigmentos extraídos destas linhagens foram identificados através da análise de cromatografia de alta eficiência como beta-caroteno e toruleno. As linhagens de leveduras mostraram potencial como microrganismos promissores para a produção comercial de carotenóides por fermentação.

Mycotoxin research in Brazil: the last decade in review

The number of research papers (128 papers) on mycotoxins published by Brazilian researchers in 1991-2000 surpassed the total number (85 papers) published in the preceding three decades (1961-1990). Thirty percent of the papers surveyed mycotoxins in foods and feeds. AFs in peanut and peanut products continued to be alarming, and high incidence and levels of FBs in corn and corn products also appeared as a serious problem. Contamination with other toxins, such as ZEA, OTA and trichothecenes, was low. Occurrence of AFM1 in milk and dairy products and patulin in apple juice needs to be verified as the results are somewhat diverging. Work on analytical methods, mycological examination and toxic effects constituted 16, 13 and 13%, respectively, of the published papers in the decade assessed. Attempts to find means of preventing/controlling fungal growth and mycotoxin production notably increased, making up 27% of the papers, including investigations on influencing factors (e.g. genotype resistance, water content/a w, relative humidity, temperature, presence of metals, type of soil, mite infestation) and antagonistic potential of other microorganisms against mycotoxin-producing fungi. Effects of plant extract, flavonoids, fungicides and other chemicals, storage bag material, adsorbents, cooking and processing of food were also studied. Thus, notwithstanding constraints on resources, Brazilian research responds to the needs of the country, reflects international concerns and recent developments in the area.

O número de artigos de pesquisa (128 artigos) sobre micotoxinas publicados por pesquisadores brasileiros em 1991-2000 superou a soma de artigos (85 artigos) publicados nas três décadas anteriores (1961-1990). Trinta por cento das publicações foi levantamento da ocorrência de micotoxinas em alimentos e rações. Aflatoxinas em amendoim e produtos de amendoim continua sendo um problema alarmante, e a alta incidência e níveis elevados de fumonisinas em milho e produtos de milho também parecem ser um problema sério. A contaminação com outras micotoxinas, como zearalenona, ocratoxina A e tricotecenos, foi baixo. A ocorrência de aflatoxina M1 em leite e laticínios e de patulina em suco de maçã precisa ser verificada, pois, há uma certa divergência nos resultados. Trabalhos sobre os métodos analíticos, estudos micológicos e efeitos tóxicos constituíram 16, 13 e 13%, respectivamente, dos artigos publicados na década avaliada. A busca de meios de prevenção/controle da contaminação fúngica e produção de micotoxinas aumentou notadamente, perfazendo 27% dos artigos, incluindo investigações sobre fatores influentes (por exemplo, resistência de genótipos, conteúdo/atividade de água, umidade relativa, temperatura, presença de metais, tipo de solo, infestação com inseto) e o potencial antagônico de outros microrganismos contra os fungos produtores de micotoxinas. Os efeitos de extrato de planta, flavonóides, fungicidas e outros químicos, sacos utilizados para estocagem, adsorventes e processamento de alimentos foram também estudados. Portanto, apesar das limitações de recursos, a pesquisa brasileira responde as necessidades do país, reflete as preocupações internacionais e os desenvolvimentos recentes na área.

Carotenoid contents in Brazilian tomato products / Teores de carotenoides em produtos de tomate

The carotenoid contents in Brazilian tomato produc0ts were previously determined at our laboratory by using raw material. The data regarding the development of new products, the advances on processing technologies and analytical techniques, need to be updated. In this context, the present study was carried out in order to determine the carotenoid contents of processed tomato products by means of HPLC. Samples of ketchup, sauce, paste, pulp and dried tomato were purchased at supermarkets in Campinas, Brazil. For each product, five different lots from each of the three brands (a total of 65 samples) were purchased, each lot consisting of three randomly collected packages. The lycopene and -carotene concentration ranges (g/g) were 188-261 and 9.3-13 for paste, 111-203 and 5.1-7.0 for ketchup, 77-117 and 4.4-7.3 for pulp, 93-112 and 5.1-6.4 for sauce, 231-471 and 7.0-25 for dried tomato, respectively. Dried tomatoes, which were analyzed for the first time, showed the highest lycopene and lutein concentrations. -carotene contents in the paste and lycopene contents in tomato paste and in ketchup samples analyzed by the present study were higher than those obtained in the previous investigation.

A composição dos carotenoides em produtos de tomate foi anteriormente determinada em nosso laboratório, utilizando-se cromatografia em coluna aberta. Em virtude da introdução de novas variedades de tomate, do desenvolvimento de novos produtos e dos avanços tecnológicos nas áreas de processamento e de técnicas analíticas, esses dados necessitavam ser atualizados. Neste contexto, no presente estudo determinou-se a composição de carotenoides em produtos de tomates por meio de técnica CLAE. As amostras de extrato, catchup, polpa, molho pronto e tomate seco foram adquiridas em supermercados em Campinas-SP. Para cada produto, foram adquiridos cinco lotes diferentes de cada uma das três marcas (no total de 65 amostras), em que cada lote foi composto por três embalagens coletadas ao acaso. As faixas de licopeno e de -caroteno total (g/g) foram, respectivamente, 188-261 e 9,3-13 para extrato, 111-203 e 5,1-7,0 para catchup, 77-117 e 4,4-7,3 para polpa, 93-112 e 5,1-6,4 para molho pronto e 231-471 e 7,0-25 para tomate seco. O tomate seco, que foi analisado pela primeira vez, apresentou os maiores teores de licopeno e luteína. Os teores de -caroteno do extrato e licopeno do extrato e catchup foram maiores nas amostras analisadas neste estudo, quando comparados com os resultados obtidos no trabalho anterior.