Yeast Culture Research Articles

Synthesis of carboxymethyl cellulose using bacterial cellulose produced by kombucha scoby (symbiotic culture of bacteria and yeast) using three different solvents

RESUMO A carboximetilcelulose de sódio (CMC) é um sal celulósico obtido por meio da alcalinização da celulose e posterior eterificação do álcali celulósico com excesso de solvente orgânico. A CMC é utilizada como agente espessante, umectante, aglutinante, entre outros. Comumente ela é sintetizada a partir da celulose vegetal ou também a partir de celulose bacteriana (CB). O objetivo deste estudo foi sintetizar a CMC utilizando CB produzida pelo scoby de kombucha, para valorizar esse resíduo, utilizando 3 diferentes solventes orgânicos, isopropanol, butan-1-ol e pentan-1-ol. Após a síntese, as CMCs foram caraterizadas por rendimento, teor de umidade, grau de substituição (GS), pH, viscosidade, espectroscopia no infravermelho com transformada de Fourier (FTIR), difratometria de raios-x (DRX), termogravimetria e termogravimetria derivada (TG/DTG). Os resultados comprovaram a síntese das CMCs e apresentaram diminuição da temperatura de degradação e da cristalinidade em comparação à CB pura. Por fim, a CMC produzida com isopropanol obteve melhores resultados em comparação à literatura, apresentado maior rendimento (197,62%), maior grau de substituição (0,26 ± 0,05), maior viscosidade (11,33 ± 0,48 cP) e menor cristalinidade (32,45%). No entanto, é necessário um estudo utilizando um número maior de amostras para que se obtenha resultados mais precisos acerca do melhor solvente.

Supercritical CO2 Extraction of Wine and Beer Yeast Residues for Sustainable Bioproduct Recovery

Abstract This paper presents a study on the use of supercritical carbon dioxide (SC-CO2) extraction to isolate yeast metabolites from beer and wine matrices effectively. A major contributor to the flavour and aroma profiles of alcoholic beverages, yeast contains a rich reservoir of bioactive compounds that can be used in various industries, including food and pharmaceuticals. Conventional extraction methods often fail to obtain comprehensive and high-quality recoveries of these valuable metabolites. In response, this study explores the use of SC-CO2, a green and tuneable solvent, to overcome the limitations of conventional extraction methods. The study investigates the influence of critical process parameters such as pressure, temperature, and extraction time on the yield and composition of extracted yeast metabolites. Advanced analytical methods (gas chromatography for the determination of fatty acids and high-performance liquid chromatography for the determination of the amino acid composition of the yeast residue left after extraction) are used in the analysis to characterize the chemical profile of the extracts. The results show that SC-CO2 extraction offers a promising alternative to extract a diverse range of yeast-derived compounds, including flavour-enhancing esters, phenolic compounds, and bioactive peptides. The optimized extraction conditions show a significant improvement in extraction efficiency compared to traditional methods. Sensory analysis reveals that beverages produced with SCCO2-extracted yeast have distinctive and desirable flavour profiles. This research not only contributes to the development of extraction technologies in the beverage industry but also opens up new opportunities for the use of yeast-derived bioactive compounds in various applications. The environmentally friendly nature of SC-CO2 extraction meets the growing demand for sustainable and clean technologies in food and beverage processing. Overall, the results of this study highlight the potential of SC-CO2 extraction as a valuable tool for improving the extraction of bioactive yeast metabolites, thereby influencing the quality and sensory properties of beer and wine.

In vivo evidence for homeostatic regulation of ribosomal protein levels in Drosophila.

The ribosome is a molecular machine essential for protein synthesis, which is composed of approximately 80 different ribosomal proteins (Rps). Studies in yeast and cell culture systems have revealed that the intracellular level of Rps is finely regulated by negative feedback mechanisms or ubiquitin-proteasome system, which prevents over- or under-abundance of Rps in the cell. However, in vivo evidence for the homeostatic regulation of intracellular Rp levels has been poor. Here, using Drosophila genetics, we show that intracellular Rp levels are regulated by proteasomal degradation of excess Rps that are not incorporated into the ribosome. By establishing an EGFP-fused Rp gene system that can monitor endogenously expressed Rp levels, we found that endogenously expressed EGFP-RpS20 or -RpL5 is eliminated from the cell when RpS20 or RpL5 is exogenously expressed. Notably, the level of endogenously expressed Hsp83, a housekeeping gene, was not affected by exogenous expression of Hsp83, suggesting that the strict negative regulation of excess protein is specific for intracellular Rps. Further analyses revealed that the maintenance of cellular Rp levels is not regulated at the transcriptional level but by proteasomal degradation of excess free Rps as a protein quality control mechanism. Our observations provide not only the in vivo evidence for the homeostatic regulation of Rp levels but also a novel genetic strategy to study in vivo regulation of intracellular Rp levels and its role in tissue homeostasis via cell competition.Key words: ribosomal protein, proteasomal degradation, Drosophila.

Impact of cooperative or competitive dynamics between the yeast Saccharomyces cerevisiae and lactobacilli on the immune response of the host.

Fungi and bacteria can be found coexisting in a wide variety of environments. The combination of their physical and molecular interactions can result in a broad range of outcomes for each partner, from competition to cooperative relationships. Most of these interactions can also be found in the human gastrointestinal tract. The gut microbiota is essential for humans, helping the assimilation of food components as well as the prevention of pathogen invasions through host immune system modulation and the production of beneficial metabolites such as short-chain fatty acids (SCFAs). Several factors, including changes in diet habits due to the progressive Westernization of the lifestyle, are linked to the onset of dysbiosis statuses that impair the correct balance of the gut environment. It is therefore crucial to explore the interactions between commensal and diet-derived microorganisms and their influence on host health. Investigating these interactions through co-cultures between human- and fermented food-derived lactobacilli and yeasts led us to understand how the strains' growth yield and their metabolic products rely on the nature and concentration of the species involved, producing either cooperative or competitive dynamics. Moreover, single cultures of yeasts and lactobacilli proved to be ideal candidates for developing immune-enhancing products, given their ability to induce trained immunity in blood-derived human monocytes in vitro. Conversely, co-cultures as well as mixtures of yeasts and lactobacilli have been shown to induce an anti-inflammatory response on the same immune cells in terms of cytokine profiles and activation surface markers, opening new possibilities in the design of probiotic and dietary therapies.

Molecular Characterization of Yeast (Saccharomyces cerevisiae) Strains by Using Molecular Marker

Molecular markers reveals polymorphism at the DNA level. Molecular characterization of Yeast (Saccharomyces cerevisiae) Strains with RAPD Molecular markers. The genetic diversity allelic variations among these strains were estimated by using RAPD molecular markers. Twenty Eight RAPD primers were screened out of them eleven primers had shown amplification of which AB1-15, 1283, OPB12, OPO4, SC1 and OPB01 these six primers showed 100% polymorphism. Out of ten wine yeast strains studied at molecular level, two strains showed maximum similarity i.e. 79 % between them viz. NCIM 3045 and 3200. Genetic diversity was analyzed based on data obtained by 11 RAPD primers. Most of the primers were found 85.71 to 100% polymorphic in nature. The diversity at molecular level was analyzed with elucidian distance of 0.40. molecular markers hold great promise due to their high efficiency, adequate accuracy, and good reproducibility

Isolation and Characterization of Palm Wine Yeasts and a Comparison of Their Dough-Raising Activity With Baker’s Yeast

Isolation and Characterization of Palm Wine Yeasts and a Comparison of Their Dough-Raising Activity With Baker’s Yeast

Bio-Dealcoholization of Wines: Can Yeast Make Lighter Wines?

The relationship between climate change and viticulture has become increasingly apparent in recent years. Rising temperatures have been a critical factor in early grape ripening. This, in turn, has led to wines with imbalanced acidity and, more importantly, higher alcohol content and pH values. Today, consumers demand high-quality and healthy products, and this trend has extended to wine consumption. Consumers prefer wines with reduced alcohol content due to the health risks associated with alcohol consumption. To meet this demand, researchers have developed modified yeast strains that reduce wine alcohol content during fermentation. These strains ferment less sugar or redirect carbon metabolism. However, their use may pose challenges, such as producing undesired secondary metabolites that can affect wine characteristics. Additionally, consumers are still divided on using genetically modified organisms (GMOs) in food and beverages. This review examines the impact of climate change on wine quality and consumer perception, taking into account new technologies used to reduce wine alcohol content or produce low-alcohol-content wines, such as low-cost techniques like bio-dealcoholization performed by non-GMO wine yeast, Saccharomyces, and non-Saccharomyces.

Substituição de virginiamicina por produtos à base de levadura (Saccharomyces cerevisiae) em dietas de bovinos: Digestibilidade e parâmetros ruminais

Abstract The aim of this study was to evaluate the effect of substituting virginiamycin with yeastbased products in high-energy diets containing monensin on the nutrient digestibility and ruminal parameters of cattle. Five crossbred Nellore × Holstein steers (initial body weight 281 kg ± 9.2 kg) were allotted to a 5 × 5 Latin square design. The diets were composed in dry matter basis of corn silage (355 g kg-1), and concentrate composed of ground corn, soybean meal, soybean hulls,mineral mix, limestone, salt and urea (645 g kg-1). The treatments were: 1 - control, 18 mg kg-1 dry matter (DM) virginiamycin (VM); 2 - enriched yeast culture (Saccharomyces cerevisiae) at 7 g animal day-1 (EYC 7); 3 - enriched yeast culture (S. cerevisiae) at 14 g animal day-1 (EYC 14); 4 - autolyzed yeast (S. cerevisiae) at 7 g animal day-1 (AY 7); and 5 - autolyzed yeast (S. cerevisiae) at 14 g animal day-1 (AY 14). Monensin (18 mg kg-1 DM) was added to the mineral mix in all treatments. No significant differences (P > 0.05) were found for DM intake (mean 8.36 kg DM), organic matter (OM) digestibility (average of 58.39%), or mean ruminal pH (mean 6.75) among treatments. Moreover, the treatment did not have a significant effect on the short-chain fatty acid or ammonia nitrogen concentrations. These results indicated the possibility of replacing virginiamycin with yeast-based products at the lowest level (7 g of enriched yeast culture or autolyzed yeast per animal day-1) in high-energy diets containing monensin.

Fermentation Kinetics and Physicochemical Characterization of Nonalcoholic Wine Produced From Cashew Apple Juice

There is an increasing demand for nonalcoholic wines owing to religious beliefs, responsible driving, harmful effects of alcohol, and strict alcohol regulatory measures. Alcohol removal methods, used for nonalcoholic wine production, have negative impact on flavour characteristics and consumer acceptance. This study seeks to determine the fermentation kinetics and physicochemical properties of nonalcoholic wine developed from cashew apple juice using cold fermentation in combination with special yeast. The extracted cashew apple must was inoculated with S. ludwigii and Z. rouxii yeast culture separately and allowed to ferment at 0°C–4°C for 7 days in a completely randomized design. The pH, total soluble solids (TSS), temperature, specific gravity, titratable, volatile, and nonvolatile acidities of the fermenting must and wine were determined using standard methods. All the yeast strains maintained comparable fermentation trends with respect to pH, TSS, specific gravity, volatile acidity, titratable acidity, and fixed acid, but with different alcohol yields of 0.89 and 0.68% for S. ludwigii and Z. rouxii, respectively. The wine produced using S. ludwigii had a lower reducing sugar (3.58 mg/L) and TSS (6.65°Brix) compared to Z. rouxii (6.23 mg/L and 7.65°Brix), respectively. However, no significant differences existed in pH, titratable acidity, specific gravity, alcohol content, and colour characteristics: brightness (L∗). The matured wine products recorded 0% alcohol content and had distinct appealing colours. The application of cold fermentation combined with special yeast in nonalcoholic wine production is promising and deserves more research to explore its full potential in meeting the increasing demand for nonalcoholic wines.

Recruiting grape‐isolated antagonistic yeasts for the sustainable bio‐management of Botrytis cinerea on grapes

AbstractBotrytis cinerea is the causative agent of grey mould disease in grapes, which was linked to significant postharvest losses. This study examined three grape‐isolated yeasts (Metschnikowia aff. fructicola, Metschnikowia pulcherrima, and Hansenispora uvarum) through in vitro and in vivo tests on detached grape berries against grey mould, as well as the elucidation of their possible mechanisms of action. The antifungal mechanism of action of yeasts was determined by the lytic enzyme activity, inhibition of spore germination, biofilm activity, iron depletion, diffusible metabolites, wound‐site colonisation, mycocin, and volatile organic compounds (VOCs) production. The highest in vitro efficacy (83.13%) was observed on M. aff. fructicola, followed by M. pulcherrima (82.10%) and H. uvarum (71.66%). Metschnikowia yeasts exhibited comparable enzyme activities, including protease, β‐1,3 glucanase, gelatinase chitinase, and cellulase, while H. uvarum had a poor enzymatic activity with chitinase and gelatinase. M. aff. fructicola showed relatively higher iron depletion activity than M. pulcherrima, while M. pulcherrima outperformed via diffusible metabolites. All yeast cultures significantly reduced spore germination by at least 86%. Overall, M. aff. fructicola exhibited the highest biocontrol activity with its iron depletion, inhibition of conidial germination, biofilm formation, VOCs, and well colonisation on grape berries. M. aff. fructicola 1‐UDM outperformed all other yeasts by significantly reducing disease incidence and lesion diameter values (93.4% and 94.3%, respectively). Remarkably, H. uvarum VOCs demonstrated potential as a biofumigant for suppressing grey mould. All yeasts are well adapted to their ecological niche to bio‐protect grapes from grey mould disease.

Study of antimicrobial activity and fermentability of the yeast Wickerhamomyces anomalus in wheat dough

The work is aimed at assessing the potential of using the yeast culture Wickerhamomyces anomalus CBS605T in bread baking. The in vitro antimicrobial activity of the yeast culture Wickerhamomyces anomalus (CBS605T) against the causative agent of potato disease, Bacillus subtilis, was studied. A rheoenzymemetric assessment of the gas-forming and gas-retaining abilities of the studied strain during fermentation in wheat dough was carried out. The study showed that the yeast culture Wickerhamomyces anomalus (CBS605T) effectively inhibits the growth of Bacillus subtilis and has higher fermentation activity than the control organism Saccharomyces cerevisiae var. boulardii (Y3925), also known for antimicrobial activity against the pathogen of potato disease.

Genetic and phenotypical diversity of Pseudomonas syringae population in the Russian Federation.

Proteobacteria comprising species of Pseudomonas syringae group cause diseases of many plants around the world. The phytopathogen has a complex taxonomic structure, which is constantly being revised due to the emergence of new molecular and biochemical diagnostic methods. Here for the first time, we describe the genetic and phenotypic diversity of 57 strains of Pseudomonas syringae isolated from affected soybeans, cereals, sunflowers, and other plants in the Russian Federation from 1950 to 2019. Genetic diversity was assessed by Multi Locus Sequence Analysis (MLSA) using fragments of the genes of glyceraldehyde-3-phosphate dehydrogenase (gapdh), the DNA-directed RNA polymerase subunit D (rpoD), gyrase (topoisomerase) B subunit (gyrB), and citrate synthase I (gltA). The synthesis of syringomycin and coronatine by bacteria was assessed by the reaction of susceptible yeast culture, seedlings of barley, tomato, and sunflower, and by presence of toxin genes confirmed by PCR test. The pathogenicity of the strains was confirmed on seedlings of dicotyledonous and monocotyledonous plants of peas, soybean, sunflowers, barley and wheat, as the most affected crops. The sensitivity of bacteria to 10 antibiotics of the main mechanisms of activity and two bactericidal commercial products was tested by standard disc method. The obtained results showed a high genetic homogeneity of the Russian population of P. syringae, which infects various agricultural crops, and an increase in the proportion of antibiotic-resistant strains over the years.

Millifluidic magnetophoresis-based chip for age-specific fractionation: evaluating the impact of age on metabolomics and gene expression in yeast.

A novel millifluidic process introduces age-based fractionation of S. pastorianus var. carlsbergensis yeast culture through magnetophoresis. Saccharomyces yeast is a model organism for aging research used in various industries. Traditional age-based cell separation methods were labor-intensive, but techniques like magnetic labeling have eased the process by being non-invasive and scalable. Our approach introduces an age-specific fractionation using a 3D-printed millfluidic chip in a two-step process, ensuring efficient cell deflection in the magnetic field and counteracting magnetic induced convection. Among various channel designs, the pinch-shaped channel proved most effective for age differentiation based on magnetically labeled bud scar numbers. Metabolomic analyses revealed changes in certain amino acids and increased NAD+ levels, suggesting metabolic shifts in aging cells. Gene expression studies further underlined these age-related metabolic changes. This innovative platform offers a high-throughput, non-invasive method for age-specific yeast cell fractionation, with potential applications in industries ranging from food and beverages to pharmaceuticals.

Multi-Strain Probiotics Enhance the Bioactivity of Cascara Kombucha during Microbial Composition-Controlled Fermentation.

Kombucha is a widely consumed fermented tea beverage with diverse health benefits. In a previous study, we demonstrated that the use of cascara as a substrate results in a special kombucha beverage with high bioactivity. Traditional kombucha fermentation using a symbiotic culture of bacteria and yeast (SCOBY) can lead to inconsistent product quality because of the lack of control over microbial composition. We successfully isolated and identified yeast and bacteria, including Saccharomyces cerevisiae, Komagataeibacter rhaeticus, and Lactobacillus brevis that are appropriate starter cultures for cascara kombucha fermentation. We also demonstrated that a supplementation with lactic acid bacteria (LAB) and a mixture of S. cerevisiae and K. rhaeticus resulted in higher total polyphenol and flavonoid content of cascara kombucha compared with the traditionally fermented product using SCOBY as the inoculum. The free radical scavenging activity, inhibitory effects on α-amylase, tyrosinase activity, and antibacterial properties of cascara kombucha were also enhanced as a result of LAB supplement. These findings provide valuable insights into the controlled microbiological composition required for the fermentation of cascara kombucha, thereby ensuring consistent quality and enhanced bioactivity of the product. Further, the use of cascara as a substrate for kombucha production not only offers various health benefits and biological effects, but also repurposes by-products from the coffee industry, which contributes to sustainable development and is eco-friendly.

OBTAINING ACTIVE DRY YEASTS BIOMASS FOR THE PRODUCTION OF PIETROASA WINES

Consumers are increasingly demanding traditional beverages such as wines that are obtained through innovative technologies and that use indigenous yeasts to preserve the concept of `terroir`. The paper presents the study performed for the exploitation the microbial diversity in the Pietroasa wine region in Romania and to obtain autochthones yeasts biomass. Yeasts were isolated from grapes and must from Feteasca regala FR) and Cabernet Sauvignon (CS) varieties cultivated at the Pietroasa Viticulture and Vinification Research and Development Station. Five yeast isolates were selected and identified by the MALDI-TOF, these belonging to the S. cerevisiae and non-Saccharomyces genera. By using these yeast isolates, active dry biomass was obtained through fermentation on a substrate based on sterilised diluted must (from the Feteasca regala and Cabernet Sauvignon varieties), synthetic sterilised media, followed by freeze-drying. Subsequently, the active dry biomass was used to obtain wines, using two types of nutrients: ET (Energyvin Thiols) and NO (Nutristart® ORG). All studied yeast isolates showed a promising potential for obtaining white wine from Feteasca regala grapes and rosé wine from the Cabernet Sauvignon grapes variety.

Screening and characterization of indigenous non-Saccharomyces cerevisiae with high enzyme activity for kiwifruit wine production

To explore the diversity and fermentation potential of non-Saccharomyces cerevisiae associated with kiwifruit, indigenous yeasts isolated from kiwifruit and natural fermentation were comprehensively analyzed. A total of 166 indigenous yeasts were isolated, of which 54 representative strains were used for subsequent enzyme activity characterization. Different colorimetric methods were used to verify the ability of these strains to secrete hydrolytic enzymes, and then six strains were selected for sequential fermentation by specific activity assay. The performance of indigenous yeasts in improving organic acids, polyphenols, volatile compounds and sensory characteristics of wines was evaluated holistically. Results indicated that most sequential fermentations exhibited significant improvements in vitamin C and polyphenols. Remarkably, the involvement of Zygosaccharomyces rouxii, Meyerozyma guilliermondii, and Pichia kudriavzevii increased the concentrations of ethyl esters, acetates and alcohols, enhancing floral and tropical fruit odors and ultimately achieving the highest overall sensory acceptability, thereby highlighting their potential in kiwifruit wine fermentation.

Interactions between Starmerella bacillaris and Saccharomyces cerevisiae during sequential fermentations influence the release of yeast mannoproteins and impact the protein stability of an unstable wine

Wine protein haze formation is a problem due to grape proteins aggregation during wine storage. The cell wall components of wine yeasts, particularly high molecular weight mannoproteins, have a protective effect against haze formation, although their involvement remains poorly understood. This study aimed at characterizing glycosylated proteins released by Starmerella bacillaris and Saccharomyces cerevisiae during single and sequential fermentations in a synthetic must, and testing their impact on wine protein stability. Mannoproteins-rich extracts from sequential fermentations showed an increase in the low MW polysaccharide fraction and, when added to an unstable wine, had a greater effect on protein stability than S. cerevisiae extracts. Shotgun proteomics approaches revealed that the identified cell wall proteins exclusively found in sequential fermentations were produced by both S. bacillaris (MKC7, ENG1) and S. cerevisiae (Bgl2p). Moreover, sequential fermentations significantly increased the expression of Scw4p and 1,3-beta-glucanosyltransferase (GAS5), produced by S. cerevisiae. Finally, some of the key proteins identified might play a positive role in increasing wine protein stability.

RSM approach to pre-treatment of lignocellulosic waste and a statistical methodology for optimizing bioethanol production

A potential substitute for a sustainable and long-term energy source is bioethanol. Although converting lignocellulose to bioethanol may be more economical, eco-friendly, and effective, it would still require extensive process-development and improvement to make biofuel economically feasible. An essential stage in the production of bioethanol is the pre-treatment of biomass to dissolve lignin and recover holocellulose. In this study, waste biomass from agro-forestry was converted into bioethanol by using various pre-treatments and selecting the one that produced the best results to optimize the process using the Classical One Factor at a Time approach (COFAT). The saccharification of selected biomass was done using in-house cultures of hyper-cellulase (Bacillus stratosphericus N12) and hyper-xylanase (B. altitudinus Kd1) producers, while the yeast cultures for fermentation of biomass i.e. Streptomyces cerevisiae and Pichia stiptis were purchased as sterile cultures. COFAT used inoculum size, fermentation pH, incubation time and temperature as optimization parameters and each of these increased the amount of bioethanol produced. Additionally, the Central Composite Design (CCD) of Response Surface Methodology (RSM) contributed to the 28.44 g/L maximum bioethanol output. The pre-treatments that separated the lignin from the holocellulose allowed it to be used in another investigation for the manufacture of nanoparticles and as a biocontrol agent. Since each component of waste biomass can be used to make useful products such as bioethanol from holocellulose and an anti-fungalbiocontrol agentfrom lignin, a comprehensive approach to bioutilization of lignocellulosic biomass has been undertaken.

Effect of protatranes on the biosynthesis of extracellular enzymes of <i>Candida ethanolica</i> BKM Y-2300 T

The study aims to explore the possibility of using synthetic biologically active compounds (protatranes) to increase the lipolytic and proteolytic activity of Candida ethanolica. Protatrane 1 (tris(2-hydroxyethyl) ammonium-4chlorophenyl-sulfanyl acetate) and protatrane 2 (tris(2-hydroxyethyl) ammonium-4-chlorophenyl-sulfonyl acetate) were added to the growth medium at trace concentrations of 1×10−6–1×10−8wt% separately and together. It was established that with the introduction to the growth medium, protatranes had a multidirectional effect on the biosynthesis of enzymes by Candida ethanolica yeast. This effect was dependent on the concentrations of these compounds, as well as on their separate or combined introduction. All the studied concentrations were found to be effective for lipase synthesis, as they improved enzyme synthesis by 1.7–8.6 times. The combined use of protatranes increased enzyme synthesis by 3.4–11.7 times. For protease formation, the combined introduction of the studied protatranes at a concentration of 10−6wt% was found to be the most effective, with enzyme synthesis reaching 184.8±7.02 U/mL in the culture broth. When co-introduced, the studied protatranes tris(2-hydroxyethyl) ammonium-4-chlorophenylsulfanyl acetate and tris(2-hydroxyethyl) ammonium-4-chlorophenyl-sulfonyl acetate can be used to increase the production efficiency of extracellular lipases and proteases by Candida ethanolica. The specific effect of protatranes was revealed to be the uneven dynamics of extracellular enzyme accumulation, manifested in the presence of sharp peaks during yeast cultivation. The reasons for this unevenness require further research.

Two-step separation strategy with mixed-mode chromatography to purify recombinant human serum albumin from Pichia pastoris culture broth

Mixed-mode chromatography (MMC) with the specially-designed ligands shows good adsorption selectivity for protein separation. In this study MMC was explored to purify recombinant human serum albumin (rHSA) from Pichia pastoris culture broth, which combined bind-elute and flow-through modes. Four mixed-mode resins were evaluated for their adsorption capacities of HSA. It was found that two home-made MMC resins showed special performances. IC-B-6FF resin with indole-3-acetic acid-cysteine ligand had high adsorption and selectivity towards HSA, and SFA-B-6FF resin with sulfapyridine ligand showed minimal binding to HSA. Therefore, a two-step MMC separation strategy was designed that IC-B-6FF was used to capture rHSA directly from yeast culture broth and then SFA-B-6FF resin was used to polish the remaining impurities by flow-through mode. The results revealed that the purity of rHSA reached 95.1% with a yield of 81.2% after the two-step separation. The removal rate of host cell proteins (HCPs) was 99.8% with final HCP level as low as 50 ppm. The results demonstrate that MMC combining bind-elute and flow-through modes has a promising potential to separate protein with high selectivity and process efficiency.