Biotransformation of Phytosterols into Androstenedione-A Technological Prospecting Study.

Androstenedione (AD) is a key intermediate in the body's steroid metabolism, used as a precursor for several steroid substances, such as testosterone, estradiol, ethinyl estradiol, testolactone, progesterone, cortisone, cortisol, prednisone, and prednisolone. The world market for AD and ADD (androstadienedione) exceeds 1000 tons per year, which stimulates the pharmaceutical industry's search for newer and cheaper raw materials to produce steroidal compounds. In light of this interest, we aimed to investigate the progress of AD biosynthesis from phytosterols by prospecting scientific articles (Scopus, Web of Science, and Google Scholar databases) and patents (USPTO database). A wide variety of articles and patents involving AD and phytosterol were found in the last few decades, resulting in 108 relevant articles (from January 2000 to December 2021) and 23 patents of interest (from January 1976 to December 2021). The separation of these documents into macro, meso, and micro categories revealed that most studies (articles) are performed in China (54.8%) and in universities (76%), while patents are mostly granted to United States companies. It also highlights the fact that AD production studies are focused on "process improvement" techniques and on possible modifications of the "microorganism" involved in biosynthesis (64 and 62 documents, respectively). The most-reported "process improvement" technique is "chemical addition" (40%), which means that the addition of solvents, surfactants, cofactors, inducers, ionic liquids, etc., can significantly increase AD production. Microbial genetic modifications stand out in the "microorganism" category because this strategy improves AD yield considerably. These documents also revealed the main aspects of AD and ADD biosynthesis: Mycolicibacterium sp. (basonym: Mycobacterium sp.) (40%) and Mycolicibacterium neoaurum (known previously as Mycobacterium neoaurum) (32%) are the most recurrent species studied. Microbial incubation temperatures can vary from 29 °C to 37 °C; incubation can last from 72 h to 14 days; the mixture is agitated at 140 to 220 rpm; vegetable oils, mainly soybean, can be used as the source of a mixture of phytosterols. In general, the results obtained in the present technological prospecting study are fundamental to mapping the possibilities of AD biosynthesis process optimization, as well as to identifying emerging technologies and methodologies in this scenario.

Butter whey and corn steep liquor as sole raw materials to obtain a bioemulsifier from Yarrowia lipolytica for food oil-in-water emulsions / Soro de manteiga e milhocina como únicas matérias primas para obter bioemulsificante de Yarrowia lipolytica para emulsões óleo-em-água alimentícias

ABSTRACT: A synthetic medium containing glucose, glycerol, yeast extract (YE), and ammonium sulfate (AS) was compared to several low-cost media in their ability to produce high emulsification index (EI). The goal was to reduce the production costs of an emulsifier with application in food oil-in-water emulsions. To this end, agro-industrial by-products were screened for bioemulsifier production from Yarrowia lipolytica. The statistical analysis showed that the EIs of media containing residual frying oil from palm oil (RFO_palm) or soybean oil (RFO_soy), residual liquid from butter production (butter whey, BWhey) or cheese production (cheese whey, CWhey), supplemented with YE and AS were similar to the EI of the synthetic medium. The replacement of YE by corn steep liquor (CSL) also resulted in similar EI, except for RFO_soy. BWhey was tested with CSL without AS and similar EI (66.8%) was detected in comparison to that of the same medium with AS (66.3%). The cell-free broth obtained after Y. lipolytica growth in BWhey+CSL was successfully used to obtain vegetable oil-in-water emulsions indicating its potential application in food products.

RESUMO: O meio sintético contendo glicose, glicerol, extrato de levedura (YE) e sulfato de amônio (AS) foi comparado a meios de baixo custo para produzir alto índice de emulsificação (EI). Para este fim, os subprodutos agroindustriais foram rastreados quanto à produção de bioemulsificante por Yarrowia lipolytica. A análise estatística mostrou que EI de meios contendo óleo de fritura residual de óleo de palma (RFO_palm) ou óleo de soja (RFO_soy), líquido residual da produção de manteiga (soro de manteiga, BWhey) ou da produção de queijo (soro de queijo, CWhey), suplementado com YE e AS foram semelhantes ao EI do meio sintético. A substituição do YE por milhocina (CSL) também resultou em EI semelhante, exceto no RFO_soy. O BWhey foi testado com CSL sem AS e EI semelhante (66,8%) foi detectado em comparação com o mesmo meio com AS (66,3%). O meio isento de células obtido após o crescimento de Y. lipolytica no meio BWhey + CSL foi utilizado com sucesso para obter emulsões de óleo vegetal em água, indicando sua potencial aplicação em produtos alimentícios.

Catalytic and physical features of a naturally immobilized Yarrowia lipolytica lipase in cell debris (LipImDebri) displaying high thermostability.

Lipase activity (337 U/g dry weight of cell debris) was detected in cell debris after ultrasound treatment of Yarrowia lipolytica cells cultivated in residual frying palm oil. It is a naturally immobilized lipase with protein content of 47%, herein called LipImDebri. This immobilized biocatalyst presents low hydrophobicity (8%), that can be increased adjusting pH and buffer type. Despite apparent intact cells, electron microscopy showed a shapeless and flat surface for LipImDebri and optical microscopy revealed no cell viability. Besides, an inferior mean diameter (3.4 mm) in relation to whole cells reveals structure modification. A high negative zeta potential value (- 33.86 mV) for pH 6 and 25 °C suggests that LipImDebri is a stable suspension in aqueous solution. Fourier Transform Infrared Spectra (FTIR) expose differences between LipImDebri and extracellular lipase extract signaling a physical interaction between enzyme and cell debris, which is possibly the reason for the high thermostability (k d = 0.246 h-1; t 1/2 = 2.82 h at 50 °C, pH 7.0). A good adjustment of LipImDebri kinetic data with Hill equation (R 2 = 0.95) exposes an allosteric behavior related to the presence of more than one lipase isoform. These features reveal that LipImDebri can be a good catalyst for industrial applications.

Investigation of mitochondrial protein expression profiles of Yarrowia lipolytica in response to citric acid production.

Nitrogen-limiting condition is essential for citric acid production by Yarrowia lipolytica. Mitochondrial protein expression profiles of Y. lipolytica IMUFRJ 50,682 cells cultivated in biomass proliferation medium (YPG medium, yeast extract, peptone and glycerol) and citric acid production medium (CA medium) were analyzed to identify differences in expressed proteins in response to medium composition. The identification of 45 proteins in mitochondria of YPG medium cells and 48 proteins in mitochondria of CA medium cells were possible with proteomic analyses. Only 11 proteins were common to both conditions, showing a different expression pattern in relation to limiting and non-limiting nitrogen conditions. For both conditions, most proteins (52%-CA medium, 46%-YPG medium) were related to energy metabolism. CA medium cells expressed more carbohydrate metabolism proteins (six proteins) then YPG medium cells (three proteins) and the opposite was detected for translation proteins.

Green (Detox) juice physicochemical properties and stabilization effect of naturals emulsifiers / Propriedades físico-químicas do suco verde (detox) e efeito de estabilização de emulsificantes naturais

ABSTRACT: Green or "detox" juice is a mixture of fruit juice with vegetables, which has been used intensively by consumers seeking for healthy food. Physicochemical properties of Green juice were accessed in the present research, which brings new insights for the use of this beverage in human diet. A total phenolic content of 2833.60 mg GAE (Gallic acid equivalent)/ g of juice and a Total Antioxidant Capacity by FRAP of 323.62 µM Fe2SO4 / g of juice and by ABTS•+ of 333.11 µM Trolox/ g of juice, indicated good antioxidant properties. Low energy and reducing sugar content indicate its use for low calorie diet, but low carbohydrate and protein content prove that Green juice cannot be used as meal replacement. The addition of a microbial biosurfactant (YlBio) and chia gel as bioemulsifiers was tested in the Green juice formulation to reduce solid decantation and increase consistency. YlBio and chia gel were able to change the Newtonian behavior of the Green juice to a Pseudoplastic behavior due to stabilization properties and also increase consistency, without the need to add synthetic stabilizers.

RESUMO: O suco verde ou "detox" é uma mistura de suco de frutas com vegetais que tem sido intensamente utilizado por consumidores que buscam alimentos saudáveis. As propriedades físico-químicas do suco verde foram avaliadas no presente trabalho, o que traz novas perspectivas para o uso dessa bebida na dieta humana. Um conteúdo fenólico total de 2833,60 mg de EAG (equivalente em ácido gálico) / g de suco) e uma capacidade antioxidante total por FRAP de 323,62 µM de Fe2SO4 / g de suco e por ABTS•+ de 333,11 µM de Trolox / g de suco, indicam boas propriedades antioxidantes. Um baixo teor de energia e açúcar redutor indica seu uso em dietas de baixa caloria, mas o baixo teor de carboidratos e proteínas prova que o suco verde não pode ser usado como substituto de refeição. A adição de um biossurfactante microbiano (YlBio) e do gel de chia no suco foi testada na formulação do suco verde, para reduzir a decantação de sólidos e aumentar a consistência. YlBio e o gel de chia foram capazes de mudar o comportamento do suco de fluido Newtoniano para um fluido pseudoplástico devido às propriedades estabilizantes, e também aumentaram a consistência do suco, sem a necessidade de adição de estabilizantes sintéticos.

Preparo de curva padrão para inativação térmica da cepa de levedura comercial Saccharomyces cerevisiae WB-06 / Standard curve preparation for thermal inactivation of yeast commercials strain Saccharomyces cerevisiae WB-06

Na produção industrial de alimentos se faz necessário a inativação de microrganismos. O tipo mais comum empregado em produtos alimentícios, especialmente em bebidas fermentadas, é a levedura Saccharomyces cerevisiae. As técnicas de inativação mais empregadas consistem no aquecimento abrupto. Tal processo térmico altera propriedades sensoriais e nutritivas do produto final, devendo ocorrer de forma controlada, otimizando o tempo de operação. Neste artigo, com fim de obter parâmetros de resistência térmica da cepa comercial de levedura S. cerevisiae WB-06 foram feitos cultivos do microrganismo em meio líquido açucarado para posterior aquecimento a 60 ºC em banho térmico. A amostragem foi feita em intervalos de até 15 minutos. O valor obtido de tempo de redução decimal (D60) de 9,80 min está de acordo com a literatura.

Análise físico-química de cervejas comerciais sem glúten / Gluten free commercial beer physical-chemical analysis

A determinação de importantes parâmetros de qualidade de cervejas foram realizados em duas cervejas comerciais sem glúten (A e B). Para a análise, as amostras foram degaseificadas em sonicador Desruptor de Célula Ultra-Sônico (UNIQUE) para posterior aferição de parâmetros físico-químicos. As análises foram realizadas em equipamento Anton Paar Beer analyzer do Instituto Federal do Rio de Janeiro. Ao final, seus resultados foram comparados com trabalhos da literatura com outras duas cervejas com glúten (C e D) em mesmo tipo de aparelho. Foram encontrados em média valores para as cervejas sem glúten inferiores a 26% para o álcool (%p/p), 8% para extrato real (%p/p), 22% para extrato original (%p/p) e 20% para grau real de fermentação (RDF %).

Patent Landscape on Structured Lipids Produced by Enzyme Technology.

BACKGROUND: Lipid modification results in several benefits for the food industry, biotechnology advances and human health. Customizing bioactive lipids is very appealing because it improves the product's nutritional quality. Lipases are sustainable biocatalysts that can be reused, show selectivity towards substrates and reactions occur in mild conditions. OBJECTIVES: We aimed at systematically searching for patents deposited worldwide, that approached the production of structured lipids by using lipases as biocatalysts. METHOD: A patent-search strategy was set up in Questel-Orbit and the search strategy adopted was based on the combination of specific keywords in the title/abstract of the documents, encompassing thoroughly the search scope. We revised all patents relating structured lipids produced by enzyme reactions and provided an overview of the main objectives of the patents describing it, as well as a view of the principal depositors, years of publication and principal countries of deposit, as a mean to access the technological landscape on the subject. RESULTS: Forty-four documents, published over the last 34 years, were retrieved. Nine main patents' objectives were found, and the two major groups are: SL with bioactive properties and/or with fatty acids (FA) esterified at specific triacylglycerol positions and SL analogous of natural lipids. China, Japan and USA were the three main patent depositors. CONCLUSION: Although the number of patents retrieved was relatively low, this review indicates that SL production aiming at improvements in nutritional/health and/or physical attributes for food enhancement is a new field, and technological interest and innovation have been increasing over the last ten years.

Study of trans-trans farnesol effect on hyphae formation by Yarrowia lipolytica.

Dimorphism is an ability of certain fungi related to its adaptation to the environment and provides a selective advantage under stress conditions and is associated to the development of human diseases. Hyphae inducing- and inhibitory-effect of farnesol on hyphae formation by the dimorphic yeast Yarrowia lipolytica was evaluated through digital image analysis. The agitation speed of the culture was the most effective hyphae inducer in comparison to bovine calf serum and N-acetylglucosamine. In low agitation system, bovine calf serum was more effective for hyphae formation inducing 57 % of hyphae transition. Farnesol inhibited hyphae formation even in low concentration (300 µM) and this effect increased with increasing concentrations. In the presence of N-acetylglucosamine, this effect was more evident in comparison to the presence of bovine calf serum, which might have protected the cells from farnesol. Digital image analysis was an important tool to evaluate this phenomenon.

Factorial design to optimize biosurfactant production by Yarrowia lipolytica.

In order to improve biosurfactant production by Yarrowia lipolytica IMUFRJ 50682, a factorial design was carried out. A 2(4) full factorial design was used to investigate the effects of nitrogen sources (urea, ammonium sulfate, yeast extract, and peptone) on maximum variation of surface tension (Delta ST) and emulsification index (EI). The best results (67.7% of EI and 20.9 mN m(-1) of Delta ST) were obtained in a medium composed of 10 g 1(-1) of ammonium sulfate and 0.5 g 1(-1) of yeast extract. Then, the effects of carbon sources (glycerol, hexadecane, olive oil, and glucose) were evaluated. The most favorable medium for biosurfactant production was composed of both glucose (4% w/v) and glycerol (2% w/v), which provided an EI of 81.3% and a Delta ST of 19.5 mN m(-1). The experimental design optimization enhanced Delta EI by 110.7% and Delta ST by 108.1% in relation to the standard process.