Açaí Flan, A Functional Food with Lacticaseibacillus rhamnosus HN001 Probiotic: Physicochemical Characteristics, Probiotic Survival, Sensory Acceptance and Consumer Perception.

Research background: Açaí berry is rich in antioxidant compounds and is therefore closely associated with beneficial health effects. In this study, we aim to investigate the potential of using Lacticaseibacillus rhamnosus HN001 as a probiotic culture on açaí flan. Experimental approach: The chemical composition, physicochemical and microbiological characteristics, and sensory acceptance during refrigerated storage (5 °C for 42 days) of flan were investigated. In addition, the consumer perception of the product was evaluated using word association when consumers were shown a photo of the product with or without the added ingredients accompanied with a brief description of the product. Results and conclusions: The flan had a suitable chemical composition, mainly carbohydrates and proteins, probiotic viability reached 8 log CFU/g in the product and 4 log CFU/g after gastrointestinal simulation, typical açaí coloration, significant antioxidant activity and high sensory acceptability. The information about the ingredients and properties of the products increased the health value and positive feelings of the consumers towards the product. Novelty and scientific contribution: Açaí flan has proven to be a suitable carrier for L. rhamnosus HN001 as a probiotic culture, further enhancing the characteristic beneficial properties of the fruit. Therefore, combining this information with marketing strategies that inform consumers about the benefits of the product can further improve its acceptance. As far as we know, this is the first study on açaí flan with added probiotic culture.

Role of Acetic Acid Bacteria in Food and Beverages.

Acetic acid bacteria (AAB) are microorganisms widely distributed in nature. Although this group is involved in the spoilage of some foods, AAB are of great industrial interest, and their functionality is still poorly understood. AAB convert ethanol, sugars and polyols into various organic acids, aldehydes and ketones via oxidative fermentation. These metabolites are produced during a succession of biochemical reactions in various fermented foods and beverages, such as vinegar, kombucha, water kefir, lambic and cocoa. Furthermore, important products such as gluconic acid and ascorbic acid precursors can be produced industrially from their metabolism. The development of new AAB-fermented fruit drinks with healthy and functional properties is an interesting niche for research and the food industry to explore, as it can meet the needs of a wide range of consumers. Exopolysaccharides such as levan and bacterial cellulose have unique properties, but they need to be produced on a larger scale to expand their applications in this area. This work emphasizes the importance and applications of AAB during the fermentation of various foods, their role in the development of new beverages as well as numerous applications of levan and bacterial cellulose.

Bacterial cellulose production by Komagataeibacter hansenii can be improved by successive batch culture.

Bacterial cellulose (BC) is a biopolymer principally synthetized by strains of the genus Komagataeibacter. However, high costs and low production yield make large-scale application difficult. The aim of this work was to evaluate the effects of successive batch culture before fermentation on the ability to increase the capacity of bacterial cellulose biosynthesis by a low-producing strain. The Komagataeibacter hansenii strain ATCC 23,769 was initially cultivated in fermentation broth for two periods of 35 or 56 days under static conditions. At the end of each period of time, they were transferred to new broth to be cultivated again (new batch culture cycle) for 35 or 56 days and carried out in parallel with a 10-day fermentation to determine the quantity of BC produced. As a result, a greater increase was observed after the end of the second and third batch cultures of 56 days (increases of 137% and 187% in relation to the nonbatch cultured strain, respectively). The produced samples presented higher crystallinity and thermal properties but lower water holding capacity. Through this work, it was concluded that the longer the batch culture time was, the greater the increase in the capacity of cellulose biosynthesis, which also depended on the number of successive batch culture cycles carried out.

Spectralprint techniques coupled with chemometric tools for vinegar classifications.

Vinegar is a versatile product used for food preservation, cooking, healthcare, and cleaning. In this study, 80 vinegar of different raw materials, aging time, and for the first time by the agronomic method of raw material cultivation were classified by spectralprint techniques with chemometrics. Datasets were obtained by proton nuclear magnetic resonance (1H NMR), Fourier transforms mid-infrared (FT-IR), near-infrared (NIR), and ultraviolet-visible (UV-vis); then evaluated by common dimension (ComDim) and partial least squares-discriminant analysis (PLS-DA). NMR with PLS-DA had the best prediction performance compared to other techniques, with accuracy values between 92.3 and 100 %, followed by FT-IR and UV-vis of 80.8 and 96.0 % and NIR between 69.2 and 84.0 %. The results indicated that the classification of vinegar according to the agronomic cultivation method is more complex than aging time or raw material. However, any of these spectralprint techniques have demonstrated that they can be used in the classification of vinegar.

Maturation of honey from Uruçú-Amarela (Melipona mondury): Metagenomics, metabolomics by NMR 1H, physicochemical and antioxidant properties.

The objective of this study was to characterize the microbiota biodiversity of Uruçú-Amarela honey through metagenomics. Furthermore, the impact of maturation temperatures (20 and 30 °C) and time (0-180 days) on the physicochemical and antioxidant properties was investigated. 1H NMR was performed to verify metabolites formed during maturation. Uruçú-Amarela honey was mainly composed by lactic acid bacteria and osmophilic yeasts of genus Zygosaccharomyces. Maturation at 30 °C led to a higher fermentation activity, resulting in greater carbohydrate consumption, ethanol formation (0.0-0.6 %) and increased acidity (34.78-45.74 meq/kg) over the 180 days. It also resulted in honey with higher brown color (a* 0.7 to 3.89, b* 17.50-25.29) and antioxidant capacity, corroborating that the maturation is a suitable preservation technique for stingless bee honey, because it does not cause negative changes as it extends the shelf life of the stingless bee honey.

Nutritional Supplementation with Amino Acids on Bacterial Cellulose Production by Komagataeibacter intermedius: Effect Analysis and Application of Response Surface Methodology.

Bacterial cellulose (BC) is a biopolymer mainly produced by acetic acid bacteria (AAB) that has several applications in the medical, pharmaceutical, and food industries. As other living organisms, AAB require sources of chemical elements and nutrients, which are essential for their multiplication and metabolite production. So, the knowledge of the nutritional needs of microorganisms that have important industrial applications is necessary for the nutrients to be supplied in the appropriate form and amount. Considering that the choice of different nutrients as nitrogen source can result in different metabolic effects, this work aimed to verify the effects of amino acid supplementation in the culture media for BC production by an AAB strain (Komagataeibacter intermedius V-05). For this, nineteen amino acids were tested, selected, and optimized through a Plackett and Burman factorial design and central composite design to determine the optimal concentrations of each required amino acid. Membranes produced under optimal conditions were characterized in relation to chemical structure and properties by X-ray diffraction (XRD), thermogravimetric analysis (TGA), differential scanning calorimetry (DSC), infrared spectroscopy (FT-IR), and hydrophilic properties. Three amino acids had a significant positive effect and were required: aspartic acid (1.5 g L-1), phenylalanine (1.5 g L-1), and serine (3.0 g L-1). Conversely, all sulfur and positively charged amino acids had a negative effect and reduced the production yield. After optimization and validation steps, a production level of 3.02 g L-1 was achieved. Membranes produced from optimized media by this strain presented lower crystallinity index but greater thermal and hydrophilic properties than those produced from standard HS medium.

Effects of adding spices with antioxidants compounds in red ale style craft beer: A simplex-centroid mixture design approach.

The growing demand for authentic products that provide sensory characteristics combined with health benefits has been the focus of current studies. This study developed a Red Ale style craft beer with spices such as turmeric (T), black pepper (P) and aroma hops (H), used isolated or in mixtures. A mixture design was employed to evaluate the total phenolic compounds and the antioxidant activity in the green and aged beers formulations. The spice extracts influenced the product's shelf-life. The addition of spices into the beers did not affect the physicochemical parameters that classify the Red Ale style, according to the hierarchical cluster analysis, except for aroma hops. A multiresponse optimization approach simultaneously maximized the antioxidant activity and the phenolic compounds in beers. The ideal formulation obtained for green beers was 25% T and 37.5% P and H; for aged beers, the formulation was 50% T, 20% P and 30% H.

Komagataeibacter intermedius V-05: An Acetic Acid Bacterium Isolated from Vinegar Industry, with High Capacity for Bacterial Cellulose Production in Soybean Molasses Medium.

RESEARCH BACKGROUND: Despite the great properties of bacterial cellulose, its manufacture is still limited due to difficulties in large-scale production. These problems are mainly related to low production yields and high overall costs of the conventional culture media normally used. To surpass these problems, it is necessary to identify new cheap and sustainable carbon sources. Thus, this work aims to isolate and select a high cellulose-producing Komagataeibacter strain from vinegar industry, and study its potential for bacterial cellulose synthesis in an industrial soybean co-product, known as soybean molasses, used as fermentation medium. EXPERIMENTAL APPROACH: One isolated strain was able to produce high amount of cellulose in the standard Hestrin-Schramm medium, so we tested its ability to produce this biopolymer in a soybean molasses medium. The characteristics and properties of the produced bacterial cellulose membranes were analyzed by thermogravimetric analysis, X-ray diffraction, infrared spectroscopy, water-holding capacity and rehydration ratio. Genetic analysis of the selected strain served to determine its genus and species. RESULTS AND CONCLUSIONS: An isolated strain that produced the highest amount of cellulose in Hestrin-Schramm medium (3.7 g/L) was genetically identified as Komagataeibacter intermedius V-05. This strain produced 10.0 g/L of cellulose in soybean molasses medium. Membranes from both substrates had similar chemical structure, crystallinity and thermal degradation. Soybean molasses proved to be a suitable alternative medium for biosynthesis of cellulose in comparison with the standard medium. In addition to providing higher production yield, the membranes showed great structural characteristics, similar to those obtained from standard medium. NOVELTY AND SCIENTIFIC CONTRIBUTION: In this research, we have isolated and identified a Komagataeibacter strain which exhibits a high capacity for cellulose production in soybean molasses. The isolation and selection of strains with high capacity for microbial metabolite production is important for decreasing bioprocess costs. Furthermore, as there is a necessity today to find cheaper carbon sources to obtain microbial products at a lower cost, soybean molasses represents an interesting alternative medium to produce bacterial cellulose for its industrial application.

Vinegar from Anacardium othonianum Rizzini using submerged fermentation.

BACKGROUND: Anacardium othonianum Rizzini is a native Cerrado fruit, recently described in the literature. Its use is restricted to its native region and there is a lack of studies regarding production of vinegar from the pulp. This work aims to investigate the production of A. othonianum Rizzini vinegar using submerged fermentation. RESULTS: The density, alcohol content, proximal composition, pH, color coordinates, and chromatographic profile of the volatile compounds were analyzed in the slurry, fermented juice, and vinegar produced from the corpulent parts of A. othonianum Rizz. Sensory acceptance and willingness to pay were also assessed with vinegar at 4% and 6% of total acidity. The results indicated compliance with European legislation and the presence of volatile compounds such as carbon dioxide, acetic acid, ethanol, and acetaldehyde in the analyzed vinegars. Our results indicate the potential of vinegar production from A. othonianum, with 74% and 86% willingness to pay. CONCLUSIONS: The process of transformation of the fruit pulp into new products can contribute to fruit valorization and consequent preservation of the plant in the Cerrado biome. To the best of our knowledge, this is the first report of volatile compounds and minerals in A. othonianum Rizz. slurry. Our observations can be used as a basis for future studies regarding the preparation of vinegars from this species and for investigating their application in cooking and guiding consumer perception. © 2020 Society of Chemical Industry.

Elaboration of mangaba vinegar by semi-solid fermentation combined with enzymatic activity: chemical characterization and sensory evaluation / Elaboração de vinagre de mangaba por fermentação semi-sólida com ação enzimática: caracterização química e avaliação sensorial

ABSTRACT: The objective of this study was to produce vinegar from mangaba pulp using semi-solid alcoholic fermentation combined with the enzymatic activity of pectinase and to investigate the chemical composition and sensory characteristics of the final product. was evaluated for volatile acidity and the reduced dry extract was evaluated for ashes, alcohol content, sulfates, pH, total phenolic compounds, total carotenoids, color parameters, yield, productivity, and sensory analysis. Average and standard deviation was used for descriptive statistics. Principal component analysis (PCA) was applied to all variables except total carotenoid content. Physicochemical characterization of the raw and alcoholically fermented pulp was also carried out. The main results showed that, in the vinegar, the reduced dry extract, volatile acidity, pH, and ashes were 44.3±1.5 (g/L), 4.4±0.1 (% w/v), 3.1±0.0, and 3.0±0.41 (g/L), respectively. The total phenolic compound content and total carotenoid content for the mangaba vinegar were 19.2±8.20. mg/100 g and 2.6±0.6. mg/100 g, respectively. The conversion yield from ethanol to acetic acid was 90%. PCA showed that pH and volatile acidity had a strong influence on the product, and there was a strong positive correlation between color and aroma. The final product met all legal requirements, showing that it is possible to produce mangaba vinegar with antioxidant potential for consumers. In the sensory evaluation, it was favored by the tasters, demonstrating potential economic value in the Cerrado fruit.

RESUMO: Objetivou-se produzir vinagre, a partir da polpa de mangaba por fermentação alcoólica semi-sólida com ação enzimática através da pectinase, investigar a composição química e avaliação sensorial do produto final. O vinagre foi avaliado através da acidez volátil, extrato seco reduzido, cinzas, teor alcoólico, sulfatos, pH, compostos fenólicos totais, carotenoides totais, parâmetros de cor, rendimento, produtividade e análise sensorial. Os dados foram submetidos a estatística descritiva com média e desvio padrão. Foi aplicado a análise de componentes principais (ACP) para todas as variáveis, exceto para análise de carotenoides totais. Também foi realizada a caracterização físico-química da polpa e fermentado alcoólico. Os principais resultados mostraram que, no vinagre, extrato seco reduzido, acidez volátil, pH e cinzas foram, respectivamente, 44,3±1,5 (g/L), 4,4±0,1 (% m/v), 3,1±0,0, 3,0±0,41 (g/L). Os compostos fenólicos totais e carotenoides totais apresentaram valores de 19,2±8,20 (mg/100 g) e 2,6±0,6 (mg/100 g), respectivamente. O rendimento de conversão de etanol a ácido acético foi de 90%. ACP foi aplicada nas variáveis físico-químicas do vinagre no qual os parâmetros de cor, pH e acidez volátil apresentaram forte influência no produto e, para os atributos da análise sensorial, cor e aroma apresentaram uma forte correlação positiva entre si. O produto final atendeu a todos os quesitos legais, demonstrando ser possível a produção de vinagre de mangaba com potencial antioxidante. Na avaliação sensorial teve boa aceitação pelos provadores, valorizando o uso deste fruto do Cerrado.

Acetic Acid Fermentation of Soybean Molasses and Characterisation of the Produced Vinegar.

Soybean molasses is a by-product from the production of protein concentrate from soybean meal that predominantly contains sugars, with sucrose as the major component. In Brazil, soybean molasses is used for animal feed or it is discarded, although some industries use it to produce ethanol. This study aims to evaluate the parameters required for the acetic acid fermentation of soybean molasses, and characterise the resultant vinegar. To study the most suitable parameters for the acetic acid fermentation, vinegar was produced from the alcohol fermentation of soybean molasses through eight fermentation cycles: five for adaptation and three for production. The average acidity of the acetic acid fermentation product was 50.60 g/L, with an acetic acid fermentation yield, total yield of acetic acid in broth and productivity 65.01%, 92.76% and 0.033 g/(L·h), respectively. The vinegar produced from soybean molasses has an acidity of 5.07% (m/V), residual ethanol content 0.17% (m/V), sugars 7.86% (m/V), dry extract 14.67% (m/V), ash 2.27% (m/V) and a density of 1.023 g/cm3. The contents of total phenolics and isoflavones decreased after the alcohol and acetic acid fermentations. Moreover, the isoflavones profile of the fermented product comprised only three forms: daidzein, glycitin and genistin. According to our results, 3460 L of vinegar can be produced for every tonne of soy molasses, with an acetic acid concentration of 40 g/L, the minimum required by the legislation on vinegar production. Thus, these findings demonstrate that soy molasses represents a useful raw material for the production of vinegar.

In vitro effects of Bifidobacterium lactis-based synbiotics on human faecal bacteria.

Synbiotic supplements contain pre- and probiotics and are used to modulate gut microbiota composition. This study aimed to investigate effects of two synbiotic mixtures on human faecal bacteria in vitro. Short chain fructooligosaccharides (FOS) (1% w/v) combined with either Bifidobacterium lactis Bb12 or Bifidobacterium lactis HN019 (106 colony-forming units (CFU)/mL)] were added to pH-controlled anaerobic batch cultures inoculated with human faeces. Maltodextrin (1% w/v), FOS (1% w/v) and the probiotic strains were also tested individually. Effects on bacteria, short-chain fatty acids (SCFAs) and branched-chain fatty acids (BCFAs) were assessed over 48 h. With maltodextrin, FOS and the synbiotic mixtures, there was a significant increase in total bacteria and bifidobacteria numbers, compared to the negative control or probiotics alone. Increases in Atopobium cluster and Clostridium coccoides-Eubacterium rectale group occurred with FOS and maltodextrin, respectively. Additionally, maltodextrin, FOS and synbiotics resulted in a greater production of acetate and butyrate (SCFAs) compared to the negative control and probiotics alone, whereas concentrations of iso-valerate (BCFA) were lower with these treatments. In conclusion, synbiotic-induced in vitro bacterial changes and changes in SCFAs concentrations were not different from those observed with FOS alone. These data suggest that metabolic effects of these synbiotics are largely driven by the prebiotic component.

Acetic Acid Bacteria in the Food Industry: Systematics, Characteristics and Applications.

The group of Gram-negative bacteria capable of oxidising ethanol to acetic acid is called acetic acid bacteria (AAB). They are widespread in nature and play an important role in the production of food and beverages, such as vinegar and kombucha. The ability to oxidise ethanol to acetic acid also allows the unwanted growth of AAB in other fermented beverages, such as wine, cider, beer and functional and soft beverages, causing an undesirable sour taste. These bacteria are also used in the production of other metabolic products, for example, gluconic acid, l-sorbose and bacterial cellulose, with potential applications in the food and biomedical industries. The classification of AAB into distinct genera has undergone several modifications over the last years, based on morphological, physiological and genetic characteristics. Therefore, this review focuses on the history of taxonomy, biochemical aspects and methods of isolation, identification and quantification of AAB, mainly related to those with important biotechnological applications.

Saccharomyces boulardii: Optimization of simultaneous saccharification and fermentation of cell production in organic and conventional apple substrate pulp.

A face-centered factorial design was used to study the influence of temperature, cellulase, and pectinase concentration on the production of Saccharomyces boulardii cells during simultaneous saccharification and fermentation of organic and conventional apple substrate pulp. The effects of the variables fermentation temperature (25-35 °C), pectinase concentration (5-25 µL/100 g), and cellulase concentration (4-8 µL/100 g) were analyzed by multiple regression and polynomial models of second order, providing the ideal conditions for yeast cultivation. Cellular production of apple substrates was expressed in log CFU/mL. The optimum condition for temperature was 27.5 °C, and 20 and 5 µL/100 g for pectinase and 8 and 7 µL/100 g for cellulase concentrations for organic and conventional apple pulp, respectively. The observed viability values were in agreement with the predicted values of 8.352 log CFU/mL (organic) and 8.317 log CFU/mL (conventional) apple pulps, thus proving the effectiveness of the models.

Behavior of Leishmania major metacyclic promastigotes during the course of infection and immune response development in resistant versus susceptible hosts

Pouco se conhece sobre os epítopos derivados de promastigotas metacíclicos de Leishmania que são importantes para a regulação ou destruição do parasita, como alvos de ação imunológica no hospedeiro vertebrado. Neste estudo, nós investigamos um método alternativo para obter promastigotas metacíclicos de Leishmania major, pela avaliação do curso da infecção e reação de hipersensibilidade do tipo retardado (HTR) em hospedeiros resistentes e susceptíveis. Promastigotas não-infectantes (procíclicos) de L. major, recentemente isolados de amastigotas, foram selecionados pela adesão a colunas de lã de vidro negativamente carregadas, enquanto que promastigotas metacíclicos não se aderem à coluna e podem ser recuperados com facilidade. Condições ótimas de cromatografia foram validadas por análise estatística. O rendimento médio de parasitas obtidos após separação em colunas de lã de vidro e a viabilidade dos promastigotas foram estimados por microscopia óptica. Os promastigotas metacíclicos tiveram um rendimento médio de 43,5% a 57,5%. Camundongos BALB/c (susceptíveis) e camundongos C57BL/6 (resistentes) apresentaram padrões distintos de lesões cutâneas, os primeiros com lesões mais agressivas, induzidas por promastigotas metacíclicos. As respostas à reação de HTR foram maiores nos grupos de camundongos C57BL/6, submetidos à infecção com promastigotas metacíclicos. Estes resultados indicam que o novo método poderia ser integrado aos protocolos existentes para estudar a metaciclogênese de parasitas do gênero Leishmania in vivo.

Behavior of Leishmania major metacyclic promastigotes during the course of infection and immune response development in resistant versus susceptible hosts

Little is known on the epitopes derived from metacyclic promastigotes of Leishmania that are important on the regulation or destruction of the parasite, as targets of immune attack in the vertebrate host. In this study we investigated an alternative method to obtain metacyclic promasigotes of Leishmania major, as evaluated by the course of infection and delayed-type hipersensitivity (DTH) in resistant versus susceptible inbred mice. Non-infective (procyclic) promastigotes of L. major recently transformed from tissue amastigotes were attached to a negatively charged glass-wool column, whereas metacyclic promastigotes were not bound to columns and could be easily recovered. Optimal chromatography conditions were validated through statistical analyses. Parasite average yield from glass wool columns and promastigote viability were estimated by light microscopy. Metacyclic promastigotes yielded 43.5% to 57.5%. Different patterns of cutaneous lesions were obtained in BALB/c (susceptible) and C57BL/6 (resistant) mice, the former with highly infective lesions induced by metacyclic promastigotes. DTH responses proved to be higher in groups of C57BL/6 mice which were infected with metacyclic promastigotes. These results indicate that the new method could be integrated with the investigation of metacyclogenesis of Leishmania in vivo.

Pouco se conhece sobre os epítopos derivados de promastigotas metacíclicos de Leishmania que são importantes para a regulação ou destruição do parasita, como alvos de ação imunológica no hospedeiro vertebrado. Neste estudo, nós investigamos um método alternativo para obter promastigotas metacíclicos de Leishmania major, pela avaliação do curso da infecção e reação de hipersensibilidade do tipo retardado (HTR) em hospedeiros resistentes e susceptíveis. Promastigotas não-infectantes (procíclicos) de L. major, recentemente isolados de amastigotas, foram selecionados pela adesão a colunas de lã de vidro negativamente carregadas, enquanto que promastigotas metacíclicos não se aderem à coluna e podem ser recuperados com facilidade. Condições ótimas de cromatografia foram validadas por análise estatística. O rendimento médio de parasitas obtidos após separação em colunas de lã de vidro e a viabilidade dos promastigotas foram estimados por microscopia óptica. Os promastigotas metacíclicos tiveram um rendimento médio de 43,5% a 57,5%. Camundongos BALB/c (susceptíveis) e camundongos C57BL/6 (resistentes) apresentaram padrões distintos de lesões cutâneas, os primeiros com lesões mais agressivas, induzidas por promastigotas metacíclicos. As respostas à reação de HTR foram maiores nos grupos de camundongos C57BL/6, submetidos à infecção com promastigotas metacíclicos. Estes resultados indicam que o novo método poderia ser integrado aos protocolos existentes para estudar a metaciclogênese de parasitas do gênero Leishmania in vivo.

Behavior of Leishmania major metacyclic promastigotes during the course of infection and immune response development in resistant versus susceptible hosts

Little is known on the epitopes derived from metacyclic promastigotes of Leishmania that are important on the regulation or destruction of the parasite, as targets of immune attack in the vertebrate host. In this study we investigated an alternative method to obtain metacyclic promasigotes of Leishmania major, as evaluated by the course of infection and delayed-type hipersensitivity (DTH) in resistant versus susceptible inbred mice. Non-infective (procyclic) promastigotes of L. major recently transformed from tissue amastigotes were attached to a negatively charged glass-wool column, whereas metacyclic promastigotes were not bound to columns and could be easily recovered. Optimal chromatography conditions were validated through statistical analyses. Parasite average yield from glass wool columns and promastigote viability were estimated by light microscopy. Metacyclic promastigotes yielded 43.5% to 57.5%. Different patterns of cutaneous lesions were obtained in BALB/c (susceptible) and C57BL/6 (resistant) mice, the former with highly infective lesions induced by metacyclic promastigotes. DTH responses proved to be higher in groups of C57BL/6 mice which were infected with metacyclic promastigotes. These results indicate that the new method could be integrated with the investigation of metacyclogenesis of Leishmania in vivo.

Pouco se conhece sobre os epítopos derivados de promastigotas metacíclicos de Leishmania que são importantes para a regulação ou destruição do parasita, como alvos de ação imunológica no hospedeiro vertebrado. Neste estudo, nós investigamos um método alternativo para obter promastigotas metacíclicos de Leishmania major, pela avaliação do curso da infecção e reação de hipersensibilidade do tipo retardado (HTR) em hospedeiros resistentes e susceptíveis. Promastigotas não-infectantes (procíclicos) de L. major, recentemente isolados de amastigotas, foram selecionados pela adesão a colunas de lã de vidro negativamente carregadas, enquanto que promastigotas metacíclicos não se aderem à coluna e podem ser recuperados com facilidade. Condições ótimas de cromatografia foram validadas por análise estatística. O rendimento médio de parasitas obtidos após separação em colunas de lã de vidro e a viabilidade dos promastigotas foram estimados por microscopia óptica. Os promastigotas metacíclicos tiveram um rendimento médio de 43,5% a 57,5%. Camundongos BALB/c (susceptíveis) e camundongos C57BL/6 (resistentes) apresentaram padrões distintos de lesões cutâneas, os primeiros com lesões mais agressivas, induzidas por promastigotas metacíclicos. As respostas à reação de HTR foram maiores nos grupos de camundongos C57BL/6, submetidos à infecção com promastigotas metacíclicos. Estes resultados indicam que o novo método poderia ser integrado aos protocolos existentes para estudar a metaciclogênese de parasitas do gênero Leishmania in vivo.