Disrupted Yeast Cells Research Articles

Characterization of yeast protein isolates extracted via high-pressure homogenization and pH shift: A promising protein source enriched with essential amino acids and branched-chain amino acids.

In the global food industry, plant-based protein isolates are gaining prominence as an alternative to animal-based counterparts. However, their nutritional value often falters due to insufficient essential amino acids. To address this issue, our study introduces a sustainable protein isolate derived from yeast cells, achieved through high-pressure homogenization (HPH) and alkali pH-shifting treatment. Subjected to HPH pressures ranging from 60 to 120MPa and 1 to 10 cycles, higher pressure and cycle numbers resulted in enhanced disruption of yeast cells. Combining HPH with alkali pH-shifting treatment significantly augmented protein extraction. Four cycles of HPH at 100MPa yielded the optimized protein content, resulting in a yeast protein isolate (YPI) with 75.3g protein per 100g powder, including 30.0g of essential amino acids and 18.4g of branched-chain amino acids per 100g protein. YPI exhibited superior water and oil-holding capacities compared to pea protein isolate, whey protein isolate (WPI), and soy protein isolate. Although YPI exhibited lower emulsifying ability than WPI, it excelled in stabilizing protein-stabilized emulsions. For foaming, YPI outperformed others in both foaming ability and stabilizing protein-based foam. In conclusion, YPI surpasses numerous plant-based protein alternatives in essential amino acids and branched-chain amino acids contents, positioning it as an excellent candidate for widespread utilization as a sustainable protein source in the food industry, owing to its exceptional nutritional advantages, as well as emulsifying and foaming properties. PRACTICAL APPLICATION: This study introduces a sustainable protein isolate derived from yeast cells. YPI exhibited considerable promise as a protein source. Nutritionally, YPI notably surpassed plant-based protein isolates in EAA and BCAA contents. Functionally, YPI demonstrated superior water-holding and oil-holding capacities, as well as an effective emulsion and foam stabilizer.

Bacterial endosymbionts of a nitrogen-fixing yeast Rhodotorula mucilaginosa JGTA-S1 - insights into a yet unknown micro-ecosystem.

Rhodotorula mucilaginosa JGTA-S1 is a yeast strain capable of fixing nitrogen and improving nitrogen nutrition in rice plants because of its nitrogen-fixing endobacteria, namely Stutzerimonas (Pseudomonas) stutzeri and Bradyrhizobium sp. To gain a deeper understanding of yeast endosymbionts, we conducted a whole-genome shotgun metagenomic analysis of JGTA-S1 cells grown under conditions of nitrogen sufficiency and deficiency. Our results showed that the endosymbiont population varied depending on the nitrogen regime. Upon mechanical disruption of yeast cells, we obtained endosymbionts in culturable form viz. Bacillus velezensis and Staphylococcus sp. under nitrogen-replete conditions and Lysinibacillus telephonicus., Brevibacillus sp., and Niallia circulans under nitrogen-depleted conditions. S. stutzeri and Bradyrhizobium sp. the previously reported endosymbionts remained unculturable. The culturable endosymbionts Staphylococcus sp. and Bacillus velezensis appear to possess genes for dissimilatory nitrate reduction (DNRA), an alternative pathway for ammonia synthesis. However, our findings suggest that these endosymbionts are facultative as they survive outside the host. The fitness of the yeast was not affected by curing of these microbes. Curing the yeast diazotrophic endosymbionts took a toll on its fitness. Our results also showed that the populations of S. stutzeri and B. velezensis increased significantly under nitrogen-depleted conditions compared to nitrogen-sufficient conditions. The importance of DNRA and nitrogen fixation is also reflected in the metagenomic reads of JGTA-S1.

Unconventional Extraction Methods of Oleaginous Yeast Cell Pretreatment and Disruption

Extraction is one of the most commonly used methods for obtaining and purifying chemical compounds for commercial usage. The aim of this study was to evaluate the effect of unconventional permeabilization and cell disruption methods on the yield of lipid extraction from cells of the oleaginous yeast Yarrowia lipolytica. Batch cultures in a medium with molasses and waste post-frying oil were carried out. The biomass was subjected to pulsed electric field (PEF), high-pressure processing (HPP), ultrasounds (US), and several conventional processing techniques with chemical and mechanical agents (glass beads, acetone, Triton and Tween surfactants). The effectiveness of the applied methods, either on cell permeabilization or cell disruption, was investigated by analyzing the oil and total protein extraction yield and oil leaching efficiency, as well as by using microscope images. The PEF and US treatments proved to be effective permeabilization methods as a step of sample pretreatment for extraction. These unconventional physical methods could efficiently increase intracellular lipid extraction yield in solvent applications.

Valorization of Spent Brewer’s Yeast Bioactive Components via an Optimized Ultrasonication Process

The increasing need for sustainable waste management and food fortification requires continuous agri-food biotechnological innovation. Spent brewer’s yeast (SBY) is a mass-produced underutilized by-product of the brewery industry and has elevated bioactive potential. The current study presents a streamlined ultrasonic SBY cell lysis method, with the main goal of bioactive compound valorization. The influence of selected ultrasonication parameters on protein release and, implicitly, on the cell disruption efficiency, was assessed. The SBY derivatives resulting from the ultrasonic cell lysis were SBY extracts (SBYEs) and cell walls (SBYCWs), which were evaluated in terms of protein content, antioxidant activity (AOA) and total polyphenol content. Scanning electron microscopy (SEM) and FT-IR spectroscopy were used to characterize SBYCWs in relation to the morphological and chemical transformations that follow ultrasonic yeast cell disruption. The optimal ultrasonication conditions of 6.25% SBY concentration, 40 °C and 33.33% duty cycle (DC) ensured the most efficient lysis. The SBY derivatives with the most elevated antioxidant activity were obtained at temperatures below 60 °C. SBYCWs had the highest polyphenol content and a relatively high content of β-glucan under these parameters. Optical microscopy and SEM confirmed the release of intracellular content and separation of SBYCWs.

Disruption of Yeast Cells Xanthophyllomyces Dendrorhous (Phaffia Rhodozyma) by Vibration Resonant Low-Frequency Cavitator

The goal of the study is to optimize the mode of disruption of the yeast Phaffia rhodozyma KNH 1 by a vibration-resonant low-frequency cavitator (VLC). The destruction of the cell biomass of yeast culture P. rhodozyma strain KNH 1 was carried out in VLC with water cooling, the capacity of 800 W, and resonant frequencies of vibrations of 30 Hz, 35 Hz, 37 Hz, 37.8 Hz, 39 Hz, 50 Hz, and in the presence of nitrogen in the reaction medium. Our data suggest that the yield of processed biomass by the treatment of yeast culture in VLC depends on the culture age and the mode of the treatment. Thus, for the six-day culture, we got the highest yield by its processing in VLC at 35 Hz for 75 min. The highest yield from the five-day culture was obtained after the treatment in VLC for 1 h at 37-37.8 Hz. The lowest yield of the disrupted yeast cells was obtained after 5 h of treatment in VLC at 37.8 Hz. The high level of yeast cell disruption can be used for the preparation of glucans aqueous solutions. Our data show that for such a level of disruption to treat five-day culture of P. rhodozyma in VLC at 37 Hz resonance frequency with nitrogen gas, bubbling through the reaction medium is economically profitable. For the first time, this study demonstrates the established optimal mode of destruction of yeast cells of P. rhodozyma strain KNH1 for the action of the vibration-resonance low-frequency cavitator or VLC. Ana¬lysis of the presented data indicates that the claimed me¬thod is convenient, efficient, and technologically justified.

High hydrostatic pressure-assisted extraction of lipids from Lipomyces starkeyi biomass.

The purpose of this study is to evaluate the effect of high hydrostatic pressure (HHP) as a novel approach for yeast cell disruption and lipid extraction from Lipomyces starkeyi DSM 70295 grown in glucose medium (40g/L and C/N:55/1) at initial pH of 5.0, 25°C, and 130rpm for 8 days. HHP extraction conditions including pressure, time, and temperature were optimized by response surface methodology. The high speed homogenizer-assisted extraction (HSH) was also used for comparison. The biomass subjected to HHP was examined under scanning electron microscopy and light microscope. A maximal lipid yield of 45.8±2.1% in dry cell basis (w/w) was achieved at 200MPa, 40°C, and 15min, while a minimum yield of 15.2±0.9% was observed at 300MPa, 40°C, and 10min (p<0.05). The lipid yield decreased with increasing pressure. It was demonstrated that low pressure (200MPa) collapsed the cells, while high pressure (400MPa) created protrusions on the cell wall and cell fragments spread in the environment. This study favors HHP as a promising method for Lipomyces oil extraction. PRACTICAL APPLICATION: Single-cell oils are considered future alternatives to plant-based oils as food additives and dietary supplements. Oleaginous microorganisms accumulate oils in their cell plasma, which makes extraction essential. One of the main obstacles with existing methods is the utilization of strong acids to destroy cell walls. This study aims to demonstrate high hydrostatic pressure as a rapid method for lipid extraction from oleaginous yeast Lipomyces starkeyi.

Lipid reduction to improve clarification and filterability during primary recovery of intracellular products in yeast lysates using exogenous lipase

AbstractBACKGROUNDThe yeast Pichia pastoris is a popular host organism for production of a range of biological products, several of which are intracellular. The disruption of yeast cells by homogenization also releases large quantities of lipids, which can foul the downstream membranes and chromatography matrices used for purification. This work examines lipid removal from yeast cells following homogenization by enzymatic degradation and its impact on the performance of the subsequent centrifugation and filtration.RESULTSLipase treatment of cell homogenate at 37 °C for 2 h, followed by clarification using a scaled‐down mimic of disc stack centrifugation, resulted in a 6.5‐fold improvement in solids removal when compared to untreated feed material. The lipase‐treated and untreated materials that had undergone initial centrifugation were then tested for filtration performance by passing the material through a 0.45 μm polyethylene sulfone membrane under constant flux. A 50% increase in throughput was observed in comparison to the untreated material.CONCLUSIONThese proof‐of‐concept data suggest enzymatic digestion of lipids, analogous to the widely performed DNA reduction using nucleases, could be a valuable process improvement strategy. © 2021 The Authors. Journal of Chemical Technology and Biotechnology published by John Wiley &amp; Sons Ltd on behalf of Society of Chemical Industry (SCI).

Uncovering the hidden bacterial ghost communities of yeast and experimental evidences demonstrates yeast as thriving hub for bacteria

Our major concern was to address “yeast endobacteria” which was based on a few reports in the recent past where bacteria may find yeast as a niche for survival. In this study, we report the microbiota of twenty-nine axenic yeast cultures recovered from different habitats based on their 16S rRNA gene-amplicon metagenomes. Yeasts were identified based on D1/D2 or ITS gene sequences. Bacterial diversity was widespread, varied and rich among all yeasts except for four strains. Taxa belonging to the phylum Firmicutes, Proteobacteria, Actinobacteria and Bacteroidetes and the genera; Streptococcus, Propionibacterium were common to all the yeasts. Candida tropicalis was used as a model organism to confirm bacteria through fluorescence in situ hybridization (FISH), isolating and re-introducing the isolated bacteria into the yeast. FISH analysis confirmed the endobacteria of C. tropicalis and we have successfully isolated four bacteria only after lysis and disruption of yeast cells. These bacteria were identified as species of Pseudomonas, Chryseobacterium, Lysinibacillus and Propionibacterium. Guestimates indicate 95% of bacterial species of C. tropicalis are yet-to-be-cultivated. We have successfully reintroduced mCherry tagged Pseudomonas into C. tropicalis. Also, auto-fluorescent Prochlorococcus and Rhodopseudomonas could be introduced into C. tropicalis while mCherry tagged E. coli or Salmonella could not be introduced. FISH analysis confirmed the presence of both native and infected bacterial cells present in C. tropicalis. Our findings unveil the insights into the ghost microbiota associated with yeast, which otherwise are considered to be axenic cultures. Their inherent occurrence, together with co-cultivation experiments under laboratory conditions suggests that yeasts are a thriving hub for bacterial communities.

Spent brewer's yeast as a source of high added value molecules: a systematic review on its characteristics, processing and potential applications.

Development of new strategies to add-value to agro-industrial by-products are of environmental and economical importance. Innovative and low-cost sources of protein and bioactive peptides have been explored worldwide. Spent brewer's yeast (SBY) is the second most relevant by-product from the brewing industry, and despite its nutritional (about 50% protein, dry weight) and technological potential, it is still underused or needs to be disposed of. SBY cells need to be disrupted to release intracellular and cell wall proteins. This procedure has been performed using autolysis, glass bead milling, enzymatic hydrolysis and ultrasound processing. Enzymatic treatment is usually performed without prior purification and is a challenging process, which involves multiple factors, but has been successfully used as a strategy to add value to agro-industrial by-products. Scope and approach: in this review, we particularly focused on enzymatic hydrolysis as a strategy to promote SBY valorisation, illustrating the state-of-the-art processes used to produce protein extracts from this material as well as exploring fundamental concepts related to the particularities of yeast cell disruption and protein hydrolysis. Furthermore, innovative applications of value-added yeast by-products in food, biotechnological and pharmaceutical industries are presented and discussed. Key findings and conclusions: the discovery of valuable compounds found in spent yeasts as well as the development of new processing methodologies have been widening the possibilities of reuse and transformation of SBY as an ingredient and innovative matrix. Once released, yeast proteins and peptides may be applied as an innovative non-animal protein source or a functional and bioactive ingredient.

Iron oxide magnetic nanoparticles exhibiting zymolyase-like lytic activity

Iron oxide (Fe3O4) magnetic nanoparticles (MNPs) were found to exhibit an intrinsic yeast lytic activity with enzyme kinetics similar to that of natural zymolyase. The Fe3O4 MNPs were synthesized by a co-precipitation method and were then employed to catalyze the disruption of yeast cells. The Fe3O4 MNPs accelerated the biocatalytic reaction to completion within 10 min. Compared to natural zymolyase, the newly discovered biocatalytic activity of Fe3O4 was outstanding and remained stable against temperature and pH changes. More importantly, Fe3O4 MNPs with excellent activity could be successfully synthesized using a high-gravity co-precipitation method promising for large scale production in industrial applications. Considering its cost-effectiveness and ease of preparation, Fe3O4 was expected to show great potential for the digestion of yeast cell walls through serving as nanomaterial-based artificial enzymes or nanozymes.

Unveiling the binding and orientation of the antimicrobial peptide Plantaricin 149 in zwitterionic and negatively charged membranes.

Antimicrobial peptides are a large group of natural compounds which present promising properties for the pharmaceutical and food industries, such as broad-spectrum activity, potential for use as natural preservatives, and reduced propensity for development of bacterial resistance. Plantaricin 149 (Pln149), isolated from Lactobacillus plantarum NRIC 149, is an intrinsically disordered peptide with the ability to inhibit bacteria from the Listeria and Staphylococcus genera, and which is capable of promoting inhibition and disruption of yeast cells. In this study, the interactions of Pln149 with model membranes composed of zwitterionic and/or anionic phospholipids were investigated using a range of biophysical techniques, including isothermal titration calorimetry, surface tension measurements, synchrotron radiation circular dichroism spectroscopy, oriented circular dichroism spectroscopy, and optical microscopy, to elucidate these peptides' mode of interactions and provide insight into their functional roles. In anionic model membranes, the binding of Pln149 to lipid bilayers is an endothermic process and induces a helical secondary structure in the peptide. The helices bind parallel to the surfaces of lipid bilayers and can promote vesicle disruption, depending on peptide concentration. Although Pln149 has relatively low affinity for zwitterionic liposomes, it is able to adsorb at their lipid interfaces, disturbing the lipid packing, assuming a similar parallel helix structure with a surface-bound orientation, and promoting an increase in the membrane surface area. Such findings can explain the intriguing inhibitory action of Pln149 in yeast cells whose cell membranes have a significant zwitterionic lipid composition.

Parametric and phenomenological studies about ultrasound-enhanced biosorption of phenolics from fruit pomace extract by waste yeast

In this work, sonication (20-kHz) was conducted to assist the biosorption of phenolics from blueberry pomace extracts by brewery waste yeast biomass. The adsorption capacity of yeast increased markedly under ultrasonic fields. After sonication at 394.2 W/L and 40 °C for 120 min, the adsorption capacity was increased by 62.7% compared with that under reciprocating shaking. An artificial neural network was used to model and visualize the effects of different parameters on yeast biosorption capacity. Both biosorption time and acoustic energy density had positive influences on yeast biosorption capacity, whereas no clear influence of temperature on biosorption process was observed. Regarding the mechanism of ultrasound-enhanced biosorption process, the amino and carboxyl groups in yeast were considered to be associated with the yeast biosorption property. Meanwhile, ultrasound promoted the decline of the structure order of yeast cells induced by phenolic uptake. The interactions between yeast cells and phenolics were also affected by the structures of phenolics. Moreover, the mass transfer process was simulated by a surface diffusional model considering the ultrasound-induced yeast cell disruption. The modeling results showed that the external mass transfer coefficient in liquid phase and the surface diffusion coefficient under sonication at 394.2 W/L and 40 °C were 128.5% and 74.3% higher than that under reciprocating shaking, respectively.

Microbial cell disruption methods for efficient release of enzyme L-asparaginase

The efficacy of a simple laboratory method for cell disruption based on the glass bead stirring, sonication, osmotic shock, freezing and grinding, or use of solvents and detergents was assessed in this study, via measurements of the release of total protein and L-asparaginase activity. Three different microbial sources of L-asparaginase were used: Escherichia coli BL21 (DE3), Leucosporidium muscorum, and Aspergillus terreus (CCT 7693). This study adjusted and identified the best procedure for each kind of microorganism. Sonication and glass bead stirring led to obtaining filamentous fungus cell-free extracts containing high concentrations of soluble proteins and specific activity; however, sonication was the best since it obtained 4.61 ± 0.12 IU mg−1 after 3 min of operation time. Mechanical methods were also the most effective for yeast cell disruption, but sonication was the technique which yielded a higher efficiency releasing 7.3 IUtotal compared to glass bead stirring releasing 2.7 IUtotal at the same operation time. For bacterium, sonication proved to be the best procedure due to getting the highest specific activity (9.01 IU mg−1) and total enzyme activity (61.7 IU). The data presented lead to conclude that the mechanical methods appeared to be the most effective for the disintegration of the all microbial cells studies. This is the first report related to the experimental comparison of L-ASNase extraction procedures from different microorganisms, which can also be used for extracting periplasm located enzymes from other organisms.

Extraction of carotenoids from Xanthophyllomyces dendrorhous using ultrasound‐assisted and chemical cell disruption methods

AbstractCarotenoids are natural pigments that can be synthesized by plants, algae, and microorganisms. Carotenoids are found intracellularly when bio‐produced by microorganisms. Thus, the cell wall rigidity makes pigment extraction difficult at times, so the application of various cell rupture and extraction techniques is made necessary. Therefore, this study aimed to perform Xanthophyllomyces dendrorhous Y‐10921 cell rupture using dimethyl sulphoxide (DMSO) and Ultrasound‐Assisted methods in order to recover carotenoids. The maximum recovery of specific carotenoids using the DMSO method was 680 μg/g associated with acetone extraction and 614 μg/g with the Ultrasound‐Assisted method (acetone:methanol, 7:3 v/v ratio). The results demonstrate that the Ultrasound‐Assisted method did not require a long extraction time, it had an easy biocompounds recovery, and furthermore, it was effective on yeast cell disruption.

Yeast cell disruption strategies for recovery of intracellular bio-active compounds — A review

Abstract Yeasts are cheap, attractive and easily available residual sources of valuable bio-active compounds. Extraction of these compounds requires to break the yeast cells. So efficient damage of cell wall has become an important issue to be resolved. The aim of this paper is to review the potential of some emerging cell disruption techniques for recovery of intracellular bio-active compounds from Baker's yeast including mechanical (bead mill, high pressure homogenization, ultrasonication), and non-mechanical (electrical, physical, chemical and enzymatic) techniques, as well as some newly developed methods. The advantages and drawbacks of different cell disruption methods were summarized by considering the energy consumption, the interaction of the disruption methods with downstream operations and the process economics of alternative strategies. Finally, some future directions for research areas are proposed. Industrial relevance Wine making process entails the generation of significant amount of waste yeast, which represents an attractive source of valuable compounds that has been relatively unexploited to date. To retain the valuable cell content, effective cell disruption strategies are needed to break the rigid yeast cell walls. This review summarizes the state of the art of some emerging cell disruption techniques for recovery of intracellular bio-active compounds from yeasts including mechanical (bead mill, high pressure homogenizer, ultrasonication), and non-mechanical (electrical, physical, chemical and enzymatic) techniques. Thereby, it identifies the process economics of alternative strategies by considering the interaction of the disruption methods with downstream operations as well as the current situations and future research needs.

Ultrasonic disruption of yeast cells: Underlying mechanism and effects of processing parameters

During the ultrasonic treatment of yeast cells, the damage to the cell walls and membranes was monitored by assessing the release of cell wall polysaccharides and intracellular proteins, respectively. At a low acoustic intensity (10 W/cm2), the polysaccharides were released faster than the proteins, whereas at higher acoustic intensities (24 and 39 W/cm2), this trend was reversed. At elevated temperatures, additional cell wall polysaccharides were released, whereas fewer intracellular proteins were released. Increasing processed volumes, initial cell concentrations, and salt concentrations led to the decrease of both fractions. However, the total releases per ultrasound treatment remained constant regardless of the processed volumes, and increased with the increase of initial cell concentrations. The results suggest that the ultrasonic disruption of yeast cells begins with the breakdown of the cell wall before continuing to the cell membrane. These findings may offer new avenues for exploring more efficient cell disruption or microbial inactivation processes. Ultrasonic technology has been intensively studied for microbial inactivation or cell disruption. It is believed that the inner cell membrane is the target of the ultrasonic damage. The mechanism of microbial inactivation via ultrasound involves the thinning of the cell membranes. In this study, we monitored the ultrasonic damage to yeast cell walls and membranes by monitoring the release of cell wall polysaccharides and intracellular proteins, respectively. Our results demonstrate that the ultrasonic disruption of yeast cells begins with the breakdown of the cell wall before continuing to the cell membrane. Increasing the temperature weakens the cell wall and thermally coagulates the intracellular proteins. Increasing the processed volumes, initial cell concentrations, and salt concentrations reduces the releases of cell wall polysaccharides and intracellular proteins from the viewpoint of yeast cells. However, the total releases of each fraction per ultrasound treatment remain constant regardless of the processed volumes, and increase with increasing initial cell concentrations. Our results may offer new insights towards the exploration of more efficient industrial processes for microbial inactivation or cell disruption, and in elucidating the effects of processing parameters.

Evaluation of the efficiency of different disruption methods on yeast cell wall preparation for β-glucan isolation.

Selected methods for yeast cell disruption were evaluated to establish their suitability for cell wall preparation in the process of β-glucan isolation. The effect of different disruption methods on contents of total saccharides, β-glucans and proteins in the produced cell walls preparations was analyzed. The degree of cell wall purification from intracellular components was established on the basis of the ratio of solubilised material. The investigated methods included: cell exposure to hot water (autoclaving), thermally-induced autolysis, homogenization in a bead mill, sonication and their combinations. Experimental systems were prepared in water (pH 5.0 and pH 7.0) and Tris-HCl buffer (pH 8.0). The Saccharomyces cerevisiae yeast cell wall preparations with the highest degree of cytosol component release and purification of β-glucans were produced by 30 min of cell homogenization with zirconium-glass beads (0.5 mm in diameter). This was confirmed by the highest ratio of solubilised material (approx. 64%–67%). The thus-produced preparations contained ca. 60% of total saccharides, 13%–14% of β(1,3)/(1,6)-glucans, and approx. 35% of crude proteins. Similar results were obtained after autolysis coupled with bead milling as well as with sonication, but the time required for these processes was more than 24 h. Homogenization in a bead mill could be valuable for general isolation procedures because allows one to eliminate the different autolytic activity of various yeast strains.

Disruption and protein release by ultrasonication of yeast cells

Abstract Protein is an important intracellular component of yeast cells with many beneficial functions to human health. In this study, disruption and protein releasing kinetics of Saccharomyces cerevisiae cells were investigated using an ultrasonic probe system. The effects of acoustic power and duty cycle of a sonicator on cell disruption and protein release were examined. The efficiency of cell disruption was evaluated by measurements of electrical conductivity, UV-spectroscopy and cell size. Higher degree of cell disruption was observed with increasing acoustic power and duty cycle. The relationship between protein release and cycle time at various process conditions was studied; and the data were fit to a first order kinetic expression. The analysis of these kinetic data led to the proposing of a simple model for protein releasing kinetics that involves acoustic power and duty cycle as parameters. Finally, different sonication systems have been compared, with the bath-type sonicator being less effective for yeast cell disruption and protein release compared to horn-type sonicator. Industrial relevance The cytoplasm of ( S . cerevisiae ) yeast cell is a rich source of bio-products (proteins, cytoplasmic enzymes, polysaccharides, etc.) valuable for the food industry. For good recovery of these intracellular bio-products, efficient breakage of the cell walls is a necessary step. In this study, disruption and protein releasing kinetics of S . cerevisiae cells were investigated using an ultrasonic probe system.

Improved cell disruption of Pichia pastoris utilizing aminopropyl magnesium phyllosilicate (AMP) clay

An efficient method for Pichia cell disruption that employs an aminopropyl magnesium phyllosilicate (AMP) clay-assisted glass beads mill is presented. AMP clay is functionalized nanocomposite resembling the talc parent structure Si8Mg6O20(OH)4 that has been proven to permeate the bacterial membrane and cause cell lysis. The recombinant capsid protein of cowpea chlorotic mottle virus (CCMV) expressed in Pichia pastoris GS115 was used as demonstration system for their ability of self-assembly into icosahedral virus-like particles (VLPs). The total protein concentration reached 4.24 mg/ml after 4 min treatment by glass beads mill combined with 0.2 % AMP clay, which was 11.2 % higher compared to glass beads mill only and the time was half shortened. The stability of purified CCMV VLPs illustrated AMP clay had no influence on virus assembly process. Considering the tiny amount added and simple approach of AMP clay, it could be a reliable method for yeast cell disruption.

Effect of Different Variables on the Efficiency of the Baker's Yeast Cell Disruption Process to Obtain Alcohol Dehydrogenase Activity

Cell disruption process of dry baker's yeast was studied in this work to obtain maximum activity of alcohol dehydrogenase (ADH). Disruption by ultrasonication, glass beads, and combination of these two methods was compared. A 1.8-fold increase of ADH activity can be achieved by combining glass beads with ultrasonication in comparison to ultrasonication. To achieve maximum volume activity of ADH, the effect of different variables on the cell disruption process was investigated (time, glass bead diameter, mass of glass beads, and ultrasound amplitude). Using the Design-Expert© software, 2⁴ factorial experimental design was performed. Two ultrasound probes were tested: MS 73 and KE 76. Optimal conditions (process variables) for cell disruption process were obtained. Optimal ADH activities after cell disruption with MS 73 and KE 76 probes were 1,890.9 and 1,531.7 U cm⁻³, respectively. Necessary ultrasonication time and ultrasound amplitude should be at the maximum values in the investigated variable range (30 min and 62 %). Bead size should be at maximum (4 mm) when using MS 73 probe and at minimum (0.3 mm) when using KE 76 probe. Partial purification of the enzyme was carried out and it was kinetically characterized using several oxidation and reduction systems.