Cells Of Yeast Saccharomyces Cerevisiae Research Articles

Application of phosphoric acid plus hydrogen peroxide pretreatment and Aspergillus species enzymatic decomposition on rice straw for bioethanol production

Cellulose is one of the most important of natural renewable sources. Cellulase enzymes convert it into oligosaccharides and glucose. These enzymes have several applications in industries and widely used in second generation of biofuel production process. The aim of this study was to use rice straw as a carbon source of Saccharomyces cerevisiae cells to produce bioethanol. Phosphoric acid plus hydrogen peroxide chemical pretreatment was performed to obtain a cellulose-rich source of rice straw. Mesophilic filamentous fungal isolates collected from cattle manure samples, the best isolates were selected based on cellulase activities at optimal reaction conditions. These isolates were identified by DNA sequencing of ITS region. Aspergillus fumigatus and A. niger showed the highest activities of endoglucanase, exoglucanase and total cellulase enzymes at temperature 35 °C, pH 5.5, and temperature 30 °C, pH 4.5, respectively. Pretreated rice straw was hydrolyzed by enzymatic solution of two fungal isolates. The obtained sugars (8.654 g/L for A. fumigatus and 7.412 g/L for A. niger hydrolysates) were fermented by Saccharomyces cerevisiae yeast cells and converted to the bioethanol. The highest amounts of ethanol were obtained from A. fumigatus and A. niger lysates after 72 h of fermentation as 11.2 g/L and 8.29 g/L, respectively.

Modification of Saccharomyces cerevisiae Cells with Metal Hexacyanoferrates for the Construction of a Yeast-Based Fuel Cell

This research presents a simple procedure for chemically modifying yeast (Saccharomyces cerevisiae) cells with nickel hexacyanoferrate (NiHCF) and ferric hexacyanoferrate, also known as Prussian blue (PB), to increase the conductivity of the yeast cell wall. Using linear sweep voltammetry, NiHCF-modified yeast and PB-modified yeast (NiHCF/yeast and PB/yeast, respectively) were found to have better cell wall conductivity in [Fe(CN)6]3− and glucose-containing phosphate-buffered solution than unmodified yeast. Spectrophotometric analysis showed that the modification of yeast cells with NiHCF had a less harmful effect on yeast cell viability than the modification of yeast cells with PB. The use of NiHCF/yeast and PB/yeast cells in the construction of a yeast-based fuel cell allowed the maximum power densities of 62.66 mW/m2 and 94.09 mW/m2 to be achieved. These values were much higher than those obtained using unmodified yeast cells (42.25 mW/m2). NiHCF/yeast and PB/yeast fuel cells were renewed by replenishing the yeast suspension in the anolyte or the FeCl3 salt in the catholyte. This allowed 77.4% and 50.1% of the initial maximum power density of the fuel cells to be achieved.

Potential Antimicrobial and Cytotoxic Activity of Caralluma indica Seed Extract.

Background: Plant-derived phytochemicals are crucial in fighting bacterial infections and in cancer therapy. Objective: This study investigates the phytochemical composition of the ethanolic extract obtained from Caralluma indica (C. indica) seeds and assesses its antimicrobial, anticancer, and antioxidant activities. Results: GC-MS analysis found 30 phytochemicals in C. indica seeds, including 5 bioactive compounds that have been shown to have antioxidant, antimicrobial, and cytotoxicity properties, through in silico evaluation. Phytochemical screening of C. indica identified and measured the phenolic compounds, providing insight into its bioactive potential and therapeutic properties. C. indica exhibited robust antioxidant capacity (DPPH, ABTS, nitric oxide, and H2O2 radical scavenging) alongside potent antimicrobial activity against oral pathogen and cytotoxicity activity on a human oral squamous carcinoma cell line (OECM-1) (EC50 of 169.35 µg/mL) and yeast cell Saccharomyces cerevisiae (215.82 µg/mL), with a selective index of 1.27. The subminimum % MBC/MFC of C. indica significantly reduced biofilm formation against oral pathogens (p < 0.05). Molecular docking studies showed a strong correlation (r = 0.862) between antifungal and anticancer targets, suggesting that the antimicrobial agents in C. indica contribute to cancer prevention mechanisms. Conclusions: These findings propose C. indica seeds as promising candidates for combating oral pathogens, inhibiting biofilm formation, and reducing the risk of oral cancer progression.

Exploring the Potential of Nyonya Kuih Residues as the Substrate for Yeast Cell Protein Production

Nyonya kuih, a popular Malaysian traditional cuisine that have short shelf life due to their high moisture and fat contents. The unsold kuihs usually ended up as food residue. Thus, kuih talam(KT) and kuih lapis (KL), two varieties of Nyonya kuih were chosen as the investigative substrates to explore the recycling potential of this food residue. Initially, the effect of fat content on the enzymatic hydrolysability of the residues to release fermentable sugar was investigated. The efficiency of lipase-pretreatment (LP), polyvinylpyrrolidone-post-treatment (PVP) and their combination in lipid removal was determined. Then, the potential of the resulting glucose-rich hydrolysates for Saccharomyces cerevisiae yeast cell biomass production was assessed, with molasses as the positive control. Results indicated that combined LP and PVP (LP-PVP) significantly (p&lt;0.05) reduced crude fat content, while increasing the total sugar content of KT and KL hydrolysates by approximately 3.4 and 26.3 times, respectively. Yeast cell biomass produced from KT and KL hydrolysates were increased by 23% and 39%, respectively after LP-PVP treatment. Despite the yeast cells’ yield (by dry cell weight) from molasses fermentation being 19% and 11% higher than those from KT and KL hydrolysates, respectively, KT and KL hydrolysates were proven superior due to the significantly higher (p&lt;0.05) crude protein content in yeast cell produced from both hydrolysates. Further study is needed to enhance yeast cell yield from the fermentation of KT and KL hydrolysates, thereby improving the overall protein yield from the fermentation process. This study provides new insight into a novel food residue recycle strategy, promoting sustainable food production aligned with SDG 12.

Integrating the in vitro antioxidant activity of Polyacrylonitrile/Polyindole composites

AbstractIn this study, five compounds belonging to the Polymer composites (PCs) family, have been prepared with different amounts of polyacrylonitrile and polyindole in the composite. The PCs have inherently bactericidal activity, owing to the nitrile and indole groups. Antioxidant activity was assessed by using reducing power activity, superoxide radical scavenging activity and hydroxyl radical scavenging activity. In our study, butylated hydroxy toluene (BHT), ascorbic acid and α‐tocopherol, which are considered synthetic standard antioxidants, were used to compare antioxidant activity. In addition, since it is known that the MDA levels resulting from the oxidation of fats in Saccharomyces cerevisiae yeast cells are inversely proportional to the antioxidant properties, it gave an idea about the in vitro antioxidant properties of the new synthesized PCs. Reducing force capacity, superoxide radical scavenging activity and hydroxyl radical scavenging activity were found to be higher at 100 μg mL−1 concentration compared to 50 μg mL−1 concentration.

The role of hexokinases in epigenetic regulation: altered hexokinase expression and chromatin stability in yeast

BackgroundHuman hexokinase 2 (HK2) plays an important role in regulating Warburg effect, which metabolizes glucose to lactate acid even in the presence of ample oxygen and provides intermediate metabolites to support cancer cell proliferation and tumor growth. HK2 overexpression has been observed in various types of cancers and targeting HK2-driven Warburg effect has been suggested as a potential cancer therapeutic strategy. Given that epigenetic enzymes utilize metabolic intermediates as substrates or co-factors to carry out post-translational modification of histones and nucleic acids modifications in cells, we hypothesized that altering HK2 expression could impact the epigenome and, consequently, chromatin stability in yeast. To test this hypothesis, we established genetic models with different yeast hexokinase 2 (HXK2) expression in Saccharomyces cerevisiae yeast cells and investigated the effect of HXK2-dependent metabolism on parental nucleosome transfer, a key DNA replication–coupled epigenetic inheritance process, and chromatin stability.ResultsBy comparing the growth of mutant yeast cells carrying single deletion of hxk1Δ, hxk2Δ, or double-loss of hxk1Δ hxk2Δ to wild-type cells, we firstly confirmed that HXK2 is the dominant HXK in yeast cell growth. Surprisingly, manipulating HXK2 expression in yeast, whether through overexpression or deletion, had only a marginal impact on parental nucleosome assembly, but a noticeable trend with decrease chromatin instability. However, targeting yeast cells with 2-deoxy-D-glucose (2-DG), a clinical glycolysis inhibitor that has been proposed as an anti-cancer treatment, significantly increased chromatin instability.ConclusionOur findings suggest that in yeast cells lacking HXK2, alternative HXKs such as HXK1 or glucokinase 1 (GLK1) play a role in supporting glycolysis at a level that adequately maintains epigenomic stability. While our study demonstrated an increase in epigenetic instability with 2-DG treatment, the observed effect seemed to occur dependent on non-glycolytic function of Hxk2. Thus, additional research is needed to identify the molecular mechanism through which 2-DG influences chromatin stability.

Study of the adjuvant properties of beta-glucans from &lt;i&gt;Saccharomyces Cerevisiae&lt;/i&gt; yeast

To increase the effectiveness and immunogenicity of modern vaccines, especially subunit ones, it is required to use adjuvants. Polysaccharides, due to their safety and biocompatibility, are desirable candidates for the creation of vaccine adjuvants. The aim of our study was to develop a method for obtaining beta-Glucans from the yeast Saccharomyces cerevisiae cell wall, and evaluate their adjuvant properties. The high purity and non-toxicity of the resulting preparation was achieved by using enzyme complexes of cellulase and protease in combination with ultrasound (22 kHz) at the purification stage. The developed scheme allows for the yield of beta-Glucans up to 2 g from 100 g of the biomass of wet cells. The adjuvant properties of beta-Glucans were studied in 50 male BALB/c mice, weighing 16–18 g. Immunization was performed twice, with a 14-day interval, intramuscularly, 200 μl per animal. The recombinant receptor-binding domain (RBD) of the surface S protein of the SARS-CoV-2 virus (Wuhan-Hu-1 and B.1.617.2 (Delta)) was used as an antigen, at a dose of 50 μg per animal. A positive control group was administered with the antigen combined with aluminum hydroxide. As a negative control, mice injected with the saline solution were used. The titers of specific antibodies in the blood sera were determined by ELISA assays. RBD (Wuhan-Hu-1 and Delta), and S protein (Wuhan-Hu-1, Delta and Omicron) were used as antigens. The titers of virus-neutralizing antibodies were measured in neutralization tests using SARS-CoV-2 virus strains Wuhan-Hu-1, Delta (B.1.617.2) and Omicron (B.1.1.529). The results of the study have shown that beta-Glucans have the ability to enhance the production of specific and virus-neutralizing antibodies in mice immunized with RBD. The titers of specific and virus neutralizing antibodies are comparable to their levels in the group immunized with RBD and Al(OH)₃. It has been found in the experiments in white outbred ICR mice that the preparation belongs to practically non-toxic substances. Therefore, it can be concluded that the use of beta-Glucans could become a preferable alternative to the conventional adjuvants based on aluminum salts, being biocompatible, biodegradable and non-toxic substances of low labor-intensive production.

The plant rhabdovirus viroporin P9 facilitates insect-mediated virus transmission in barley.

Potassium (K+) plays crucial roles in both plant development and immunity. However, the function of K+ in plant-virus interactions remains largely unknown. Here, we utilized Barley yellow striate mosaic virus (BYSMV), an insect-transmitted plant cytorhabdovirus, to investigate the interplay between viral infection and plant K+ homeostasis. The BYSMV accessory P9 protein exhibits viroporin activity by enhancing membrane permeability in Escherichia coli. Additionally, P9 increases K+ uptake in yeast (Saccharomyces cerevisiae) cells, which is disrupted by a point mutation of glycine 14 to threonine (P9G14T). Furthermore, BYSMV P9 forms oligomers and targets to both the viral envelope and the plant membrane. Based on the recombinant BYSMV-GFP (BYGFP) virus, a P9-deleted mutant (BYGFPΔP9) was rescued and demonstrated infectivity within individual plant cells of Nicotiana benthamiana and insect vectors. However, BYGFPΔP9 failed to infect barley plants after transmission by insect vectors. Furthermore, infection of barley plants was severely impaired for BYGFP-P9G14T lacking P9 K+ channel activity. In vitro assays demonstrate that K+ facilitates virion disassembly and the release of genome RNA for viral mRNA transcription. Altogether, our results show that the K+ channel activity of viroporins is conserved in plant cytorhabdoviruses and plays crucial roles in insect-mediated virus transmission.

Reduction of Ferric Chloride in Yeast Growth Media, by Sugars and Aluminum

Iron compounds can be used in antimicrobial applications by exploiting the toxicity of divalent iron to living organisms due to the Fenton reaction. In this study, the growth inhibitory effects of ferrous sulfate FeSO4·7H2O and ferric chloride FeCl3·6H2O were observed on Metschnikowia clade and Saccharomyces cerevisiae yeast cells. The relatively high amount of reduced Fe3+ to Fe2+ in the growth medium determined by Mössbauer spectroscopy may contribute to the antimicrobial activity of ferric chloride. In order to test the reducing ability of sugars in the growth media of yeasts, the reaction of ferric chloride FeCl3·6H2O with sugars was investigated. In mixtures of FeCl3·6H2O and fructose, approximately two thirds of Fe3+ can be reduced to Fe2+. When the mixture of FeCl3·6H2O and fructose is placed on the surface of aluminum foil, an iron film is formed on the surface of the aluminum due to the reduction by both fructose and aluminum. The relative amount of Fe3+ which was reduced to Fe0 reached 68%.

Virtually identical does not mean exactly identical: Discrepancy in energy metabolism between glucose and fructose fermentation influences the reproductive potential of yeast cells

The physiological efficiency of cells largely depends on the possibility of metabolic adaptations to changing conditions, especially on the availability of nutrients. Central carbon metabolism has an essential role in cellular function. In most cells is based on glucose, which is the primary energy source, provides the carbon skeleton for the biosynthesis of important cell macromolecules, and acts as a signaling molecule. The metabolic flux between pathways of carbon metabolism such as glycolysis, pentose phosphate pathway, and mitochondrial oxidative phosphorylation is dynamically adjusted by specific cellular economics responding to extracellular conditions and intracellular demands. Using Saccharomyces cerevisiae yeast cells and potentially similar fermentable carbon sources i.e. glucose and fructose we analyzed the parameters concerning the metabolic status of the cells and connected with them alteration in cell reproductive potential. Those parameters were related to the specific metabolic network: the hexose uptake - glycolysis and activity of the cAMP/PKA pathway - pentose phosphate pathway and biosynthetic capacities - the oxidative respiration and energy generation. The results showed that yeast cells growing in a fructose medium slightly increased metabolism redirection toward respiratory activity, which decreased pentose phosphate pathway activity and cellular biosynthetic capabilities. These differences between the fermentative metabolism of glucose and fructose, lead to long-term effects, manifested by changes in the maximum reproductive potential of cells.

Abscisic acid root-to-shoot translocation by transporter AtABCG25 mediates stomatal movements in Arabidopsis.

The phytohormone abscisic acid (ABA) plays a central role in regulating stomatal movements under drought conditions. The root-derived peptide CLAVATA3/EMBRYO SURROUNDING REGION-RELATED 25 (CLE25) moves from the root to shoot for activating ABA biosynthesis under drought conditions. However, the root-to-shoot translocation of root-derived ABA and its regulation of stomatal movements in the shoot remain to be clarified. Here, we reveal that the ABA transporter ATP-binding cassette subfamily G member 25 (AtABCG25) mediates root-to-shoot translocation of ABA and ABA-glucosyl ester (ABA-GE) in Arabidopsis (Arabidopsis thaliana). Isotope-labeled ABA tracer experiments and hormone quantification in xylem sap showed that the root-to-shoot translocation of ABA and ABA-GE was substantially impaired in the atabcg25 mutant under nondrought and drought conditions. However, the contents of ABA and ABA-GE in the leaves were lower in the atabcg25 mutant than in the wild type (WT) under nondrought but similar under drought conditions. Consistently, the stomatal closure was suppressed in the atabcg25 mutant under nondrought but not under drought conditions. The transporter activity assays showed that AtABCG25 directly exported ABA and ABA-GE in planta and in yeast (Saccharomyces cerevisiae) cells. Thus, we proposed a working model in which root-derived ABA transported by AtABCG25 via xylem mediates stomatal movements in the shoot under nondrought conditions but might exhibit little effect on stomatal movements under drought conditions. These findings extend the functions of AtABCG25 and provide insights into the long-distance translocation of ABA and its role in stomatal movements.

Encapsulation and characterization of catechin and epicatechin microcapsules using yeast cell biocarriers

In this study, catechin and epicatechin compounds were encapsulated using Saccharomyces cerevisiae yeast cell and fabricated yeast cell microcarriers were subjected to bioactive analysis, conformational and morphological characterization. Encapsulation efficiency values of the yeast cell microcarriers were 24.74 and 31.87% for the epicatechin and catechin loaded yeast cell microcapsules, respectively. Total flavonoid content of the epicatechin and catechin loaded yeast cell microcapsules were determined to be 79.67 and 61.86 mg CE/g while the antiradical activity of the samples was in the range of 4.16-39.24%. FT-IR and XRD spectrum revealed that the catechin and epicatechin were encapsulated by the yeasts cell successfully.

Identification of Aly1 and Aly2 as Modulators of Cytoplasmic pH in Saccharomyces cerevisiae.

The regulation of intracellular pH in yeast (Saccharomyces cerevisiae) cells is critical for cell function and viability. In yeast, protons (H+) can be excreted from the cell by plasma membrane ATPase PMA1 and pumped into vacuoles by vacuolar H+-ATPase. Because PMA1 is critical to the survival of yeast cells, it is unknown whether other compensatory components are involved in pH homeostasis in the absence of PMA1. To elucidate how intracellular pH is regulated independently of PMA1, we employed a screening approach by exposing the yeast haploid deletion mutant library (ver 4.0) to the selective plant plasma membrane H+-ATPase inhibitor PS-1, which we previously reported. After repeated screenings and verification, we identified two proteins, Aly1 and Aly2, that play a role in the regulation of intracellular pH when PMA1 is deficient. Our research uncovers a new perspective on the regulation of intracellular pH related to PMA1 and also preliminarily reveals a role for Aly1 and Aly2 in the regulation of intracellular pH.

Chalcone derivatives disrupt cell membrane integrity of Saccharomyces cerevisiae cells and alter their biochemical composition.

Fungal infections can be serious or life threatening in severe cases, and the need to discover and find novel antifungal agents persists. Chalcones are plant-derived aromatic compounds that have been appealing synthons for pharmaceutical industry as they have good anticancer, antibacterial, antifungal and anti-inflammatory properties. Although there are few structure-activity relationship studies on chalcones, studies that link the structural features of these compounds to their mode of action are scant. Thus, in this study, we aim to clarify the relationship between chalcone derivatives and their cellular target within the yeast cell Saccharomyces cerevisiae. We observed that some chalcone compounds lead to disruption of cell membrane and cause ion leakage out of the cell. Moreover, chalcones alter the biochemical composition of yeast cells detectable by FTIR spectroscopy and bind to the DNA as shown by our titration experiments based on UV-Vis absorbance spectroscopy. Thus, their interaction with the DNA may be the major impact of these compounds on yeast cells.

Development of Simple Fluorescence Analytical Strategy for the Detection of Triazophos Using Greenish-Yellow Emissive Carbon Dots Derived from Curcuma longa.

This study describes a new method for synthesizing water-soluble carbon dots (CDs) using "Curcuma longa" (green source) named CL-CDs via a single-step hydrothermal process. The as-synthesized CL-CDs exhibited greenish-yellow fluorescence at 548nm upon excitation at 440nm. It shows good water stability and exhibits a quantum yield of 19.4%. The developed probe is utilized for sensing triazophos (TZP) pesticide via a dynamic quenching mechanism, exhibiting favorable linearity ranging from 0.5-500μM with a limit of detection of 0.0042μM. The as-prepared CL-CDs probe was sensitive and selective towards TZP. Lastly, the successful application of the CL-CDs-based fluorescent probe in water and rice samples highlights its potential as a reliable and efficient method for the detection of TZP in various real sample matrices. Eventually, bioimaging and biocompatibility aspects of CL-CDs have been assessed on Saccharomyces cerevisiae (yeast) cell and lung cancer (A549) cell lines, respectively.

Comparison of radiation protection effects between epigallocatechin gallate and ascorbic acid

Ionizing radiation can originate from naturally occurring radiation sources on the earth or it can be from man-made sources. When interacting with cells and living organisms, ionizing radiation produces free radicals, impacting biological molecules such as proteins, lipids, and DNA in the cell nucleus and membrane, leading to cell death or causing cell mutations. Epigallocathecin gallate (EGCG) and ascorbic acid (AA) are well-known natural antioxidants that have been studied and applied as potential radical scavengers. In this study, the radiation protection effects in the presence of EGCG and AA via the scavenging process of free radicals (mainly hydroxyl radicals) were examined. Saccharomyces cerevisiae yeast cells were grown in YDP liquid medium containing yeast extract, peptone, and dextrose/glucose that supplemented with EGCG and AA at different concentrations. Then, the cell cultures were irradiated with both low (gamma) and high (helium ion beam) linear energy transfer (LET) radiations to evaluate the radiation effect on the survival of the yeast cell. Both of EGCG and AA play effectively important roles as radiation-protective agent for yeast cells and the effectiveness in radiation protection of EGCG and AA at the same concentration was almost the same between these two additives.

Single Infrared Spectrum Enables Simultaneous Identification of (Bio)Chemical Nature and Particle Size of Microorganisms and Synthetic Microplastic Beads.

Populations of nearly identical chemical and biological microparticles include the synthetic microbeads used in cosmetic, biomedical, agri-food, and pharmaceutical industries as well as the class of living microorganisms such as yeast, pollen, and biological cells. Herein, we identify simultaneously the size and chemical nature of spherical microparticle populations with diameters larger than 1 μm. Our analysis relies on the extraction of both physical and chemical signatures from the same optical spectrum recorded using attenuated total reflection (ATR)-Fourier transform infrared (FTIR) spectroscopy. These signatures are the spectral resonances caused by the microparticles, which depend on their size and the absorption peaks revealing their chemical nature. We validate the method first on separated and mixed groups of spherical microplastic particles of two different diameters, where the method is used to calculate the diameter of the microspherical particles. Then, we apply the method to correctly identify and measure the diameter of Saccharomyces cerevisiae yeast cells. Theoretical simulations to help in understanding the effect of size distribution and dispersion support our results.

Cellular Stress Impact on Yeast Activity in Biotechnological Processes-A Short Overview.

The importance of Saccharomyces cerevisiae yeast cells is known worldwide, as they are the most used microorganisms in biotechnology for bioethanol and biofuel production. Also, they are analyzed and studied for their similar internal biochemical processes to human cells, for a better understanding of cell aging and response to cell stressors. The special ability of S. cerevisiae cells to develop in both aerobic and anaerobic conditions makes this microorganism a viable model to study the transformations and the way in which cellular metabolism is directed to face the stress conditions due to environmental changes. Thus, this review will emphasize the effects of oxidative, ethanol, and osmotic stress and also the physiological and genetic response of stress mitigation in yeast cells.

Dual-Pulsed Laser Ablation of Oyster Shell Producing Novel Thin Layers Deposed to Saccharomyces cerevisiae.

Dual-pulsed (DPL) laser deposition using oyster shells as targets was studied in order to find out if this method can replace the use of high-power pulsed lasers. Aspects related to changes in the morphological structure of the thin layer but also to the chemical composition of the obtained thin layer were analyzed and compared with the target as well as with the thin layers obtained with a higher power pulsed laser in a single-pulsed (SPL) regime. Orthorhombic structures were noticed with Scanning Electron Microscopy for the thin film obtained in DPL mode compared to the irregular particles obtained in SPL mode. The deacetylation process during ablation was evidenced by Fourier Transform Infrared spectroscopy, resulting in chitosan-based thin films. The effect of the obtained thin films of chitosan on the cells of baker's yeast (Saccharomyces cerevisiae) was studied. Restoration of the yeast paste into initial yeast was noticed mainly when the hemp fabric was used as support for the coating with yeas which was after that coated with chitosan thin film produced by DPL method.

Continuous Flow Separation of Live and Dead Cells Using Gravity Sedimentation.

The separation of target cell species is an important step for various biomedical applications ranging from single cell studies to drug testing and cell-based therapies. The purity of cell solutions is critical for therapeutic application. For example, dead cells and debris can negatively affect the efficacy of cell-based therapies. This study presents a cost-effective method for the continuous separation of live and dead cells using a 3D resin-printed microfluidic device. Saccharomyces cerevisiae yeast cells are used for cell separation experiments. Both numerical and experimental studies are presented to show the effectiveness of the presented device for the isolation of dead cells from cell solutions. The experimental results show that the 3D-printed microfluidic device successfully separates live and dead cells based on density differences. Separation efficiencies of over 95% are achieved at optimum flow rates, resulting in purer cell populations in the outlets. This study highlights the simplicity, cost-effectiveness, and potential applications of the 3D-printed microfluidic device for cell separation. The implementation of 3D printing technology in microfluidics holds promise for advancing the field and enabling the production of customized devices for biomedical applications.