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BioTech, Volume 12, Issue 1 (March 2023) – 24 articles

Cover Story (view full-size image): Cryopreservation and lyophilization are effective biotechnological conservation processes used in several pharmaceutical and medical fields, notably for high-quality biological-based therapies. Both biotechnological tools are successfully used for the long-term storage of biological samples and products. These processes deal with extremely low temperatures and multiple physical states of water, a universal and essential molecule for many biological lifeforms. Many natural biological examples of extreme cold and desiccation tolerance exist in animal, plant, or fungal species. The discussed similarities confirm the interest in transposing natural processes and phenomena to controlled laboratory and industrial settings in order to gain better control and modulation capacities over the metabolic activities of complex biological organisms. View this paper
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17 pages, 2288 KiB  
Article
Optimization of Melanin Production by Streptomyces antibioticus NRRL B-1701 Using Arthrospira (Spirulina) platensis Residues Hydrolysates as Low-Cost L-tyrosine Supplement
by Oranit Kraseasintra, Sritip Sensupa, Kanjana Mahanil, Sada Yoosathaporn, Jeeraporn Pekkoh, Sirasit Srinuanpan, Wasu Pathom-aree and Chayakorn Pumas
BioTech 2023, 12(1), 24; https://doi.org/10.3390/biotech12010024 - 20 Mar 2023
Cited by 1 | Viewed by 3010
Abstract
Melanin is a functional pigment that is used in various products. It can be produced by Streptomyces antibioticus NRRL B-1701 when supplemented with L-tyrosine. Arthrospira (Spirulina) platensis is a cyanobacterium with high protein content, including the protein phycocyanin (PC). During PC’s extraction, biomass [...] Read more.
Melanin is a functional pigment that is used in various products. It can be produced by Streptomyces antibioticus NRRL B-1701 when supplemented with L-tyrosine. Arthrospira (Spirulina) platensis is a cyanobacterium with high protein content, including the protein phycocyanin (PC). During PC’s extraction, biomass residues are generated, and these residues still contain various amino acids, especially L-tyrosine, which can be used as a low-cost supplement for melanin production. Thus, this study employed a hydrolysate of A. platensis biomass residue for L-tyrosine substitution. The effects of two drying methods, namely, lyophilization and dying via a hot air oven, on the proximate composition and content of L-tyrosine in the biomass residue were evaluated. The highest L-tyrosine (0.268 g L-tyrosine/100 g dried biomass) concentration was obtained from a hot-air-oven-dried biomass residue hydrolysate (HAO-DBRH). The HAO-DBRH was then used as a low-cost L-tyrosine supplement for maximizing melanin production, which was optimized by the response surface methodology (RSM) through central composite design (CCD). Using the RSM–CCD, the maximum level of melanin production achieved was 0.24 g/L, which is approximately four times higher than it was before optimization. This result suggests that A. platensis residue hydrolysate could be an economically feasible and low-cost alternative source of L-tyrosine for the production of melanin. Full article
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12 pages, 6659 KiB  
Article
A Thermostable Lipase Isolated from Brevibacillus thermoruber Strain 7 Degrades Ɛ-Polycaprolactone
by Nikolina Atanasova, Tsvetelina Paunova-Krasteva, Margarita Kambourova and Ivanka Boyadzhieva
BioTech 2023, 12(1), 23; https://doi.org/10.3390/biotech12010023 - 14 Mar 2023
Cited by 6 | Viewed by 2125
Abstract
The tremendous problem with plastic waste accumulation has determined an interest in biodegradation by effective degraders and their enzymes, such as thermophilic enzymes, which are characterized by high catalytic rates, thermostability, and optimum temperatures close to the melting points of some plastics. In [...] Read more.
The tremendous problem with plastic waste accumulation has determined an interest in biodegradation by effective degraders and their enzymes, such as thermophilic enzymes, which are characterized by high catalytic rates, thermostability, and optimum temperatures close to the melting points of some plastics. In the present work, we report on the ability of a thermophilic lipase, by Brevibacillus thermoruber strain 7, to degrade Ɛ-polycaprolactone (PCL), as well as the enzyme purification, the characterization of its physicochemical properties, the product degradation, and its disruptive effect on the PCL surface. The pure enzyme showed the highest reported optimum temperature at 55 °C and a pH of 7.5, while its half-life at 60 °C was more than five hours. Its substrate specificity referred the enzyme to the subgroup of lipases in the esterase group. A strong inhibitory effect was observed by detergents, inhibitors, and Fe3+ while Ca2+ enhanced its activity. The monomer Ɛ-caprolactone was a main product of the enzyme degradation. Similar elution profiles of the products received after treatment with ultra-concentrate and pure enzyme were observed. The significant changes in PCL appearance comprising the formation of shallower or deeper in-folds were observed after a week of incubation. The valuable enzyme properties of the lipase from Brevibacillus thermoruber strain 7, which caused a comparatively quick degradation of PCL, suggests further possible exploration of the enzyme for effective and environment-friendly degradation of PCL wastes in the area of thermal basins, or in thermophilic remediation processes. Full article
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12 pages, 1007 KiB  
Article
Large-Scale Production of Isocitric Acid Using Yarrowia lipolytica Yeast with Further Down-Stream Purification
by Svetlana V. Kamzolova, Vladimir A. Samoilenko, Julia N. Lunina and Igor G. Morgunov
BioTech 2023, 12(1), 22; https://doi.org/10.3390/biotech12010022 - 13 Mar 2023
Viewed by 2111
Abstract
Isocitric acid (ICA) refers to a group of promising regulators of energy metabolism which has antistress, antihypoxic, and antioxidant activities. In this paper, we reported a process of ICA production from rapeseed oil using yeast Yarrowia lipolytica VKM Y-2373 in a 500-L fermentor. [...] Read more.
Isocitric acid (ICA) refers to a group of promising regulators of energy metabolism which has antistress, antihypoxic, and antioxidant activities. In this paper, we reported a process of ICA production from rapeseed oil using yeast Yarrowia lipolytica VKM Y-2373 in a 500-L fermentor. The producer synthesized 64.1 g/L ICA with a product yield of 0.72 g/g and a productivity 0.54 g/L·h. We also developed an effective purification method, including a cell separation, clarification, concentration, acidification, and crystallization process, which resulted in the formation of the crystals of monopotassium salt of ICA with a purity of 99.0–99.9%. To the best of our knowledge, this is the first report on an ICA production process at an upscaled bioreactor level. Full article
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11 pages, 1772 KiB  
Article
Production, Kinetic/Thermodynamic Study, and Evaluation of the Influence of Static Magnetic Field on Kinetic Parameters of β-Fructofuranosidase from Aspergillus tamarii Kita UCP 1279 Produced by Solid-State Fermentation
by Rodrigo Lira de Oliveira, Aldeci França Araújo dos Santos, Bianca Alencar Cardoso, Thayanne Samille da Silva Santos, Galba Maria de Campos-Takaki, Tatiana Souza Porto and Camila Souza Porto
BioTech 2023, 12(1), 21; https://doi.org/10.3390/biotech12010021 - 03 Mar 2023
Cited by 1 | Viewed by 1660
Abstract
β-fructofuranosidases (FFases) are enzymes involved in sucrose hydrolysis and can be used in the production of invert sugar and fructo-oligosaccharides (FOS). This last is an important prebiotic extensively used in the food industry. In the present study, the FFase production by Aspergillus tamarii [...] Read more.
β-fructofuranosidases (FFases) are enzymes involved in sucrose hydrolysis and can be used in the production of invert sugar and fructo-oligosaccharides (FOS). This last is an important prebiotic extensively used in the food industry. In the present study, the FFase production by Aspergillus tamarii Kita UCP 1279 was assessed by solid-state fermentation using a mixture of wheat and soy brans as substrate. The FFase presents optimum pH and temperature at 5.0–7.0 and 60 °C, respectively. According to the kinetic/thermodynamic study, the FFase was relatively stable at 50 °C, a temperature frequently used in industrial FOS synthesis, using sucrose as substrate, evidenced by the parameters half-life (115.52 min) and D-value (383.76 min) and confirmed by thermodynamic parameters evaluated. The influence of static magnetic field with a 1450 G magnetic flux density presented a positive impact on FFase kinetic parameters evidenced by an increase of affinity of enzyme by substrate after exposition, observed by a decrease of 149.70 to 81.73 mM on Km. The results obtained indicate that FFases present suitable characteristics for further use in food industry applications. Moreover, the positive influence of a magnetic field is an indicator for further developments of bioprocesses with the presence of a magnetic field. Full article
(This article belongs to the Section Agricultural and Food Biotechnology)
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15 pages, 6107 KiB  
Article
Development and Evaluation of Chitosan-Based Food Coatings for Exotic Fruit Preservation
by Juan Camilo Zárate-Moreno, Diana Marcela Escobar-Sierra and Rigoberto Ríos-Estepa
BioTech 2023, 12(1), 20; https://doi.org/10.3390/biotech12010020 - 13 Feb 2023
Cited by 3 | Viewed by 2914
Abstract
Chitosan has gained agro-industrial interest due to its potential applications in food preservation. In this work, chitosan applications for exotic fruit coating, using feijoa as a case of study, were evaluated. For this, we synthetized and characterized chitosan from shrimp shells and tested [...] Read more.
Chitosan has gained agro-industrial interest due to its potential applications in food preservation. In this work, chitosan applications for exotic fruit coating, using feijoa as a case of study, were evaluated. For this, we synthetized and characterized chitosan from shrimp shells and tested its performance. Chemical formulations for coating preparation using chitosan were proposed and tested. Mechanical properties, porosity, permeability, and fungal and bactericidal characteristics were used to verify the potential application of the film in the protection of fruits. The results indicated that synthetized chitosan has comparable properties to commercial chitosan (deacetylation degree > 82%), and, for the case of feijoa, the chitosan coating achieved significant reduction of microorganisms and fungal growth (0 UFC/mL for sample 3). Further, membrane permeability allowed oxygen exchange suitable for fruit freshness and natural physiological weight loss, thus delaying oxidative degradation and prolonging shelf-life. Chitosan’s characteristic of a permeable film proved to be a promising alternative for the protection and extension of the freshness of post-harvest exotic fruits. Full article
(This article belongs to the Section Agricultural and Food Biotechnology)
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14 pages, 2371 KiB  
Article
Fabrication of a Polycaprolactone/Chitosan Nanofibrous Scaffold Loaded with Nigella sativa Extract for Biomedical Applications
by Qasim Shakir Kahdim, Najmeddine Abdelmoula, Hassan Al-Karagoly, Salim Albukhaty and Jabbar Al-Saaidi
BioTech 2023, 12(1), 19; https://doi.org/10.3390/biotech12010019 - 12 Feb 2023
Cited by 11 | Viewed by 3089
Abstract
In this study, biocompatible electrospun nanofiber scaffolds were produced using poly(-caprolactone (PCL)/chitosan (CS) and Nigella sativa (NS) seed extract, and their potential for biomedical applications was investigated. Scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), total porosity measurements, and water contact angle [...] Read more.
In this study, biocompatible electrospun nanofiber scaffolds were produced using poly(-caprolactone (PCL)/chitosan (CS) and Nigella sativa (NS) seed extract, and their potential for biomedical applications was investigated. Scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), total porosity measurements, and water contact angle measurements were used to evaluate the electrospun nanofibrous mats. Additionally, the antibacterial activities of Escherichia coli and Staphylococcus aureus were investigated, as well as cell cytotoxicity and antioxidant activity, using MTT and DPPH assays, respectively. The obtained PCL/CS/NS nanofiber mat was observed by SEM to have a homogeneous and bead-free morphology, with average diameters of 81.19 ± 4.38 nm. Contact angle measurements showed that the wettability of the electrospun PCL/Cs fiber mats decreased with the incorporation of NS when compared to the PCL/CS nanofiber mats. Efficient antibacterial activity against S. aureus and E. coli was displayed, and an in vitro cytotoxic assay demonstrated that the normal murine fibroblast cell line (L929 cells) remained viable after 24, 48, and 72 h following direct contact with the produced electrospun fiber mats. The results suggest that the PCL/CS/NS hydrophilic structure and the densely interconnected porous design are biocompatible materials, with the potential to treat and prevent microbial wound infections. Full article
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11 pages, 1577 KiB  
Article
In Vitro Anti-HIV-1 Activity of Chitosan Oligomers N-Conjugated with Asparagine and Glutamine
by Fatih Karadeniz
BioTech 2023, 12(1), 18; https://doi.org/10.3390/biotech12010018 - 08 Feb 2023
Cited by 6 | Viewed by 1906
Abstract
Chitosan oligomers (COS) are polysaccharides obtained by the hydrolyzation of chitosan. They are water-soluble, biodegradable, and have a wide range of beneficial properties for human health. Studies have shown that COS and its derivatives possess antitumor, antibacterial, antifungal, and antiviral activities. The goal [...] Read more.
Chitosan oligomers (COS) are polysaccharides obtained by the hydrolyzation of chitosan. They are water-soluble, biodegradable, and have a wide range of beneficial properties for human health. Studies have shown that COS and its derivatives possess antitumor, antibacterial, antifungal, and antiviral activities. The goal of the current study was to investigate the anti-human immunodeficiency virus-1 (HIV-1) potential of amino acid-conjugated COS compared to COS itself. The HIV-1 inhibitory effects of asparagine-conjugated (COS-N) and glutamine-conjugated (COS-Q) COS were evaluated by their ability to protect C8166 CD4+ human T cell lines from HIV-1 infection and infection-mediated death. The results show that the presence of COS-N and COS-Q was able to prevent cells from HIV-1-induced lysis. Additionally, p24 viral protein production was observed to be suppressed in COS conjugate-treated cells compared to COS-treated and untreated groups. However, the protective effect of COS conjugates diminished by delayed treatment indicated an early stage inhibitory effect. COS-N and COS-Q did not show any inhibitory effect on the activities of HIV-1 reverse transcriptase and protease enzyme. The results suggest that COS-N and COS-Q possess an HIV-1 entry inhibition activity compared to COS and further studies to develop different peptide and amino acid conjugates containing N and Q amino acids might yield more effective compounds to battle HIV-1 infection. Full article
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18 pages, 1137 KiB  
Review
Heterologous Expression of Recombinant Human Cytochrome P450 (CYP) in Escherichia coli: N-Terminal Modification, Expression, Isolation, Purification, and Reconstitution
by Tao Shang, Chee Mun Fang, Chin Eng Ong and Yan Pan
BioTech 2023, 12(1), 17; https://doi.org/10.3390/biotech12010017 - 07 Feb 2023
Cited by 2 | Viewed by 3693
Abstract
Cytochrome P450 (CYP) enzymes play important roles in metabolising endogenous and xenobiotic substances. Characterisations of human CYP proteins have been advanced with the rapid development of molecular technology that allows heterologous expression of human CYPs. Among several hosts, bacteria systems such as Escherichia [...] Read more.
Cytochrome P450 (CYP) enzymes play important roles in metabolising endogenous and xenobiotic substances. Characterisations of human CYP proteins have been advanced with the rapid development of molecular technology that allows heterologous expression of human CYPs. Among several hosts, bacteria systems such as Escherichia coli (E. coli) have been widely used thanks to their ease of use, high level of protein yields, and affordable maintenance costs. However, the levels of expression in E. coli reported in the literature sometimes differ significantly. This paper aims to review several contributing factors, including N-terminal modifications, co-expression with a chaperon, selections of vectors and E. coli strains, bacteria culture and protein expression conditions, bacteria membrane preparations, CYP protein solubilizations, CYP protein purifications, and reconstitution of CYP catalytic systems. The common factors that would most likely lead to high expression of CYPs were identified and summarised. Nevertheless, each factor may still require careful evaluation for individual CYP isoforms to achieve a maximal expression level and catalytic activity. Recombinant E. coli systems have been evidenced as a useful tool in obtaining the ideal level of human CYP proteins, which ultimately allows for subsequent characterisations of structures and functions. Full article
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13 pages, 1814 KiB  
Article
Separating and Purifying Mycosporine-like Amino Acids from Cyanobacteria for Application in Commercial Sunscreen Formulations
by Valeria Candelo and Carole Anne Llewellyn
BioTech 2023, 12(1), 16; https://doi.org/10.3390/biotech12010016 - 03 Feb 2023
Cited by 2 | Viewed by 2305
Abstract
Using algal-derived mycosporine-like amino acids (MAAs) in sunscreen formulations is constrained by low cellular concentrations of MAAs and by the high costs associated with harvesting algal cells and extracting the MAAs. Here, we report an industrial scalable method using a membrane filtration approach [...] Read more.
Using algal-derived mycosporine-like amino acids (MAAs) in sunscreen formulations is constrained by low cellular concentrations of MAAs and by the high costs associated with harvesting algal cells and extracting the MAAs. Here, we report an industrial scalable method using a membrane filtration approach to purify and concentrate aqueous extracts of MAAs. The method includes an additional biorefinery step enabling purification of phycocyanin, an established valuable natural product. Cultivated cells of the cyanobacterium Chlorogloeopsis fritschii (PCC 6912) were concentrated and homogenised to produce a feed for sequential processing through three membranes of decreasing pore size to obtain a retentate and permeate for each step. Microfiltration (0.2 µm) was used to remove cell debris. Ultrafiltration (10,000 Da) was used to remove large molecules and recover phycocyanin. Finally, nanofiltration (300–400 Da) was used to remove water and other small molecules. Permeate and retentate were analysed using UV-visible spectrophotometry and HPLC. The initial homogenised feed had a shinorine concentration of 5.6 ± 07 mg L−1. The final nanofiltered retentate resulted in a 3.3 times-purified concentrate (shinorine concentration of 18.71 ± 0.29 mg L−1). Significant process losses (35%) highlight scope for improvement. Results confirm the potential of membrane filtration to purify and concentrate aqueous solutions of MAAs with simultaneous separation of phycocyanin highlighting a biorefinery approach. Full article
(This article belongs to the Section Industrial Biotechnology)
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17 pages, 22698 KiB  
Commentary
Industrial Biotechnology Conservation Processes: Similarities with Natural Long-Term Preservation of Biological Organisms
by Alexis Laurent, Corinne Scaletta, Philippe Abdel-Sayed, Wassim Raffoul, Nathalie Hirt-Burri and Lee Ann Applegate
BioTech 2023, 12(1), 15; https://doi.org/10.3390/biotech12010015 - 31 Jan 2023
Cited by 1 | Viewed by 2156
Abstract
Cryopreservation and lyophilization processes are widely used for conservation purposes in the pharmaceutical, biotechnological, and food industries or in medical transplantation. Such processes deal with extremely low temperatures (e.g., −196 °C) and multiple physical states of water, a universal and essential molecule for [...] Read more.
Cryopreservation and lyophilization processes are widely used for conservation purposes in the pharmaceutical, biotechnological, and food industries or in medical transplantation. Such processes deal with extremely low temperatures (e.g., −196 °C) and multiple physical states of water, a universal and essential molecule for many biological lifeforms. This study firstly considers the controlled laboratory/industrial artificial conditions used to favor specific water phase transitions during cellular material cryopreservation and lyophilization under the Swiss progenitor cell transplantation program. Both biotechnological tools are successfully used for the long-term storage of biological samples and products, with reversible quasi-arrest of metabolic activities (e.g., cryogenic storage in liquid nitrogen). Secondly, similarities are outlined between such artificial localized environment modifications and some natural ecological niches known to favor metabolic rate modifications (e.g., cryptobiosis) in biological organisms. Specifically, examples of survival to extreme physical parameters by small multi-cellular animals (e.g., tardigrades) are discussed, opening further considerations about the possibility to reversibly slow or temporarily arrest the metabolic activity rates of defined complex organisms in controlled conditions. Key examples of biological organism adaptation capabilities to extreme environmental parameters finally enabled a discussion about the emergence of early primordial biological lifeforms, from natural biotechnology and evolutionary points of view. Overall, the provided examples/similarities confirm the interest in further transposing natural processes and phenomena to controlled laboratory settings with the ultimate goal of gaining better control and modulation capacities over the metabolic activities of complex biological organisms. Full article
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15 pages, 1351 KiB  
Review
Cell Immortalization: In Vivo Molecular Bases and In Vitro Techniques for Obtention
by Javier Curi de Bardet, Celeste Ramírez Cardentey, Belkis López González, Deanira Patrone, Idania Lores Mulet, Dario Siniscalco and María de los Angeles Robinson-Agramonte
BioTech 2023, 12(1), 14; https://doi.org/10.3390/biotech12010014 - 28 Jan 2023
Cited by 2 | Viewed by 3733
Abstract
Somatic human cells can divide a finite number of times, a phenomenon known as the Hayflick limit. It is based on the progressive erosion of the telomeric ends each time the cell completes a replicative cycle. Given this problem, researchers need cell lines [...] Read more.
Somatic human cells can divide a finite number of times, a phenomenon known as the Hayflick limit. It is based on the progressive erosion of the telomeric ends each time the cell completes a replicative cycle. Given this problem, researchers need cell lines that do not enter the senescence phase after a certain number of divisions. In this way, more lasting studies can be carried out over time and avoid the tedious work involved in performing cell passes to fresh media. However, some cells have a high replicative potential, such as embryonic stem cells and cancer cells. To accomplish this, these cells express the enzyme telomerase or activate the mechanisms of alternative telomere elongation, which favors the maintenance of the length of their stable telomeres. Researchers have been able to develop cell immortalization technology by studying the cellular and molecular bases of both mechanisms and the genes involved in the control of the cell cycle. Through it, cells with infinite replicative capacity are obtained. To obtain them, viral oncogenes/oncoproteins, myc genes, ectopic expression of telomerase, and the manipulation of genes that regulate the cell cycle, such as p53 and Rb, have been used. Full article
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21 pages, 6621 KiB  
Article
A Synergistic pH-Responsive Serum Albumin-Based Drug Delivery System Loaded with Doxorubicin and Pentacyclic Triterpene Betulinic Acid for Potential Treatment of NSCLC
by Zally Torres-Martinez, Daraishka Pérez, Grace Torres, Sthephanie Estrada, Clarissa Correa, Natasha Mederos, Kimberly Velazquez, Betzaida Castillo, Kai Griebenow and Yamixa Delgado
BioTech 2023, 12(1), 13; https://doi.org/10.3390/biotech12010013 - 26 Jan 2023
Cited by 3 | Viewed by 2470
Abstract
Nanosized drug delivery systems (DDS) have been studied as a novel strategy against cancer due to their potential to simultaneously decrease drug inactivation and systemic toxicity and increase passive and/or active drug accumulation within the tumor(s). Triterpenes are plant-derived compounds with interesting therapeutic [...] Read more.
Nanosized drug delivery systems (DDS) have been studied as a novel strategy against cancer due to their potential to simultaneously decrease drug inactivation and systemic toxicity and increase passive and/or active drug accumulation within the tumor(s). Triterpenes are plant-derived compounds with interesting therapeutic properties. Betulinic acid (BeA) is a pentacyclic triterpene that has great cytotoxic activity against different cancer types. Herein, we developed a nanosized protein-based DDS of bovine serum albumin (BSA) as the drug carrier combining two compounds, doxorubicin (Dox) and the triterpene BeA, using an oil-water-like micro-emulsion method. We used spectrophotometric assays to determine protein and drug concentrations in the DDS. The biophysical properties of these DDS were characterized using dynamic light scattering (DLS) and circular dichroism (CD) spectroscopy, confirming nanoparticle (NP) formation and drug loading into the protein structure, respectively. The encapsulation efficiency was 77% for Dox and 18% for BeA. More than 50% of both drugs were released within 24 h at pH 6.8, while less drug was released at pH 7.4 in this period. Co-incubation viability assays of Dox and BeA alone for 24 h demonstrated synergistic cytotoxic activity in the low μM range against non-small-cell lung carcinoma (NSCLC) A549 cells. Viability assays of the BSA-(Dox+BeA) DDS demonstrated a higher synergistic cytotoxic activity than the two drugs with no carrier. Moreover, confocal microscopy analysis confirmed the cellular internalization of the DDS and the accumulation of the Dox in the nucleus. We determined the mechanism of action of the BSA-(Dox+BeA) DDS, confirming S-phase cell cycle arrest, DNA damage, caspase cascade activation, and downregulation of epidermal growth factor receptor (EGFR) expression. This DDS has the potential to synergistically maximize the therapeutic effect of Dox and diminish chemoresistance induced by EGFR expression using a natural triterpene against NSCLC. Full article
(This article belongs to the Section Medical Biotechnology)
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13 pages, 873 KiB  
Article
Varietal Differences in Juice, Pomace and Root Biochemical Characteristics of Four Rhubarb (Rheum rhabarbarum L.) Cultivars
by Viktor Kharchenko, Nadezhda Golubkina, Alessio Tallarita, Maria Bogachuk, Helene Kekina, Anastasia Moldovan, Vladimir Tereshonok, Marina Antoshkina, Olga Kosheleva, Sergey Nadezhkin and Gianluca Caruso
BioTech 2023, 12(1), 12; https://doi.org/10.3390/biotech12010012 - 19 Jan 2023
Cited by 1 | Viewed by 1923
Abstract
The complex evaluation of varietal biochemical differences in rhubarb juice, pomace and roots is highly useful to develop an efficient processing technology. Research was carried out to compare four rhubarb cultivars (Malakhit, Krupnochereshkovy, Upryamets and Zaryanka) in terms of the quality and antioxidant [...] Read more.
The complex evaluation of varietal biochemical differences in rhubarb juice, pomace and roots is highly useful to develop an efficient processing technology. Research was carried out to compare four rhubarb cultivars (Malakhit, Krupnochereshkovy, Upryamets and Zaryanka) in terms of the quality and antioxidant parameters of juice, pomace and roots. The laboratory analyses showed a high juice yield (75–82%) with a relatively high content of ascorbic acid (125–164 mg L−1) and other organic acids (16–21 g L−1). Citric, oxalic and succinic acids accounted for 98% of the total acids amount. The juice of the cultivar Upryamets demonstrated high levels of the natural preservatives sorbic (36.2 mg L−1) and benzoic acids (11.7 mg L−1), which are highly valuable in juice production. The juice pomace proved to be an excellent source of pectin and dietary fiber, whose concentrations reached 21–24% and 59–64%, respectively. The total antioxidant activity decreased according to the following sequence: root pulp (161–232 mg GAE g−1 d.w.) > root peel (115–170 mg GAE g−1 d.w.) > juice pomace (28.3–34.4 mg GAE g−1 d.w.) > juice (4.4–7.6 mg GAE g−1 f.w.), suggesting that root pulp is a highly valuable antioxidant source. The results of this research highlight the interesting prospects of the complex rhubarb plant processing for the production of juice, containing a wide spectrum of organic acids and natural stabilizers (sorbic and benzoic acids), dietary fiber and pectin (juice pomace) and natural antioxidants (roots). Full article
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2 pages, 234 KiB  
Editorial
Acknowledgment to the Reviewers of BioTech in 2022
by BioTech Editorial Office
BioTech 2023, 12(1), 11; https://doi.org/10.3390/biotech12010011 - 17 Jan 2023
Viewed by 1472
Abstract
High-quality academic publishing is built on rigorous peer review [...] Full article
19 pages, 2863 KiB  
Article
High-Level Production of Soluble Cross-Reacting Material 197 in Escherichia coli Cytoplasm Due to Fine Tuning of the Target Gene’s mRNA Structure
by Yulia Alexandrovna Khodak, Alexandra Yurievna Ryazanova, Ivan Ivanovich Vorobiev, Alexander Leonidovich Kovalchuk, Nikolay Nikolaevich Ovechko and Petr Gennadievich Aparin
BioTech 2023, 12(1), 9; https://doi.org/10.3390/biotech12010009 - 11 Jan 2023
Cited by 3 | Viewed by 3170
Abstract
Cross-reacting material 197 (CRM197) is a non-toxic mutant of the diphtheria toxin and is widely used as a carrier protein in conjugate vaccines. This protein was first obtained from the supernatant of the mutant Corynebacterium diphtheriae strain. This pathogenic bacteria strain is characterized [...] Read more.
Cross-reacting material 197 (CRM197) is a non-toxic mutant of the diphtheria toxin and is widely used as a carrier protein in conjugate vaccines. This protein was first obtained from the supernatant of the mutant Corynebacterium diphtheriae strain. This pathogenic bacteria strain is characterized by a slow growth rate and a relatively low target protein yield, resulting in high production costs for CRM197. Many attempts have been made to establish high-yield protocols for the heterologous expression of recombinant CRM197 in different host organisms. In the present work, a novel CRM197-producing Escherichia coli strain was constructed. The target protein was expressed in the cytoplasm of SHuffle T7 E. coli cells without any additional tags and with a single potential mutation—an additional Met [−1]. The fine tuning of the mRNA structure (the disruption of the single hairpin in the start codon area) was sufficient to increase the CRM197 expression level several times, resulting in 150–270 mg/L (1.1–2.0 mg/g wet biomass) yields of pure CRM197 protein. Besides the high yield, the advantages of the obtained expression system include the absence of the necessity of CRM197 refolding or tag removal. Thus, an extensive analysis of the mRNA structure and the removal of the unwanted hairpins in the 5′ area may significantly improve the target protein expression rate. Full article
(This article belongs to the Section Medical Biotechnology)
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15 pages, 488 KiB  
Review
Recent Developments on the Performance of Algal Bioreactors for CO2 Removal: Focusing on the Light Intensity and Photoperiods
by Zarook Shareefdeen, Ali Elkamel and Zaeem Bin Babar
BioTech 2023, 12(1), 10; https://doi.org/10.3390/biotech12010010 - 11 Jan 2023
Cited by 7 | Viewed by 5082
Abstract
This work presents recent developments of algal bioreactors used for CO2 removal and the factors affecting the reactor performance. The main focus of the study is on light intensity and photoperiods. The role of algae in CO2 removal, types of algal [...] Read more.
This work presents recent developments of algal bioreactors used for CO2 removal and the factors affecting the reactor performance. The main focus of the study is on light intensity and photoperiods. The role of algae in CO2 removal, types of algal species used in bioreactors and conventional types of bioreactors including tubular bioreactor, vertical airlift reactor, bubble column reactor, flat panel or plate reactor, stirred tank reactor and specific type bioreactors such as hollow fibre membrane and disk photobioreactors etc. are discussed in details with respect to utilization of light. The effects of light intensity, light incident, photoinhibition, light provision arrangements and photoperiod on the performance of algal bioreactors for CO2 removal are also discussed. Efficient operation of algal photobioreactors cannot be achieved without the improvement in the utilization of incident light intensity and photoperiods. The readers may find this article has a much broader significance as algae is not only limited to removal or sequestration of CO2 but also it is used in a number of commercial applications including in energy (biofuel), nutritional and food sectors. Full article
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13 pages, 1531 KiB  
Article
Formulation of a Simulated Wastewater Influent Composition for Use in the Research of Technologies for Managing Wastewaters Generated during Manned Long-Term Space Exploration and Other Similar Situations—Literature-Based Composition Development
by Bimi Shrestha, Rafael Hernandez, Dhan Lord B. Fortela, Wayne Sharp, Andrei Chistoserdov, Daniel Gang, Emmanuel Revellame, William E. Holmes and Mark E. Zappi
BioTech 2023, 12(1), 8; https://doi.org/10.3390/biotech12010008 - 10 Jan 2023
Viewed by 2359
Abstract
The prospect of humans inhabiting planetary bodies is gaining interest among research and development communities, with the moon being considered as a transitory base camp and Mars the next planet humans will inhabit. NASA’s Mission to Mars program is set to have humans [...] Read more.
The prospect of humans inhabiting planetary bodies is gaining interest among research and development communities, with the moon being considered as a transitory base camp and Mars the next planet humans will inhabit. NASA’s Mission to Mars program is set to have humans inhabiting Mars within on-planet space camps by the Year 2030, which has tremendously increased research and development for space exploration—including research oriented toward human life support in long-term planetary lodging camps. The sustenance of human life on Mars will not be trivial due to the unavailability of an appropriate atmosphere and usable water. This situation requires a self-sustaining human life support system that can provide the basic needs such are breathable air, potable water, food, and energy. The feasibility of sending a payload with resources adequate to support long-term human inhabitation is not reasonable, which means every resource within a Mars space camp is valuable, including human-produced wastes. A biorefinery system that treats wastewater and can also produce valuable products such as oxygen, food, and energy offers a form of circular utilization of valuable resources. To conduct research for such systems requires a wastewater influent that is representative of the wastewater to be generated by the space crew within this isolated, confined environment, which is different from what is generated on Earth due to limited variability in diet, human activity, and lifestyle in this confined area. Collection of actual wastewater influent from an isolated environment supporting humans is challenging. Additionally, to ensure a safe working environment in the laboratory and avoid the imposed threat of handling actual human feces, the proposed synthetic, non-human feces containing wastewater influent formulation offers an easy-to-produce and safer-to-handle option. This paper reviews several synthetic wastewater compositions that have been formulated for space exploration purposes. None of the formulations were found to be realistic nor adequate for a space-camp-type scenario. Thus, the formulation of a synthetic wastewater for simulating a wastewater influent from a human space-based camp is proposed in this paper. In addition, the physical, chemical, and biodegradation characteristics of the final formulation designed are presented to illustrate the value of the proposed influent formulation. Full article
(This article belongs to the Topic Advances in Environmental Biotechnology (AEB))
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12 pages, 1366 KiB  
Article
An Optimized CoBRA Method for the Microfluidic Electrophoresis Detection of Breast Cancer Associated RASSF1 Methylation
by Claire Aibel, Adriana Coll De Peña and Anubhav Tripathi
BioTech 2023, 12(1), 7; https://doi.org/10.3390/biotech12010007 - 09 Jan 2023
Cited by 4 | Viewed by 2432
Abstract
Although breast cancer screening assays exist, many are inaccessible and have high turnaround times, leaving a significant need for better alternatives. Hypermethylation of tumor suppressor genes is a common epigenetic marker of breast cancer. Methylation tends to occur most frequently in the promoter [...] Read more.
Although breast cancer screening assays exist, many are inaccessible and have high turnaround times, leaving a significant need for better alternatives. Hypermethylation of tumor suppressor genes is a common epigenetic marker of breast cancer. Methylation tends to occur most frequently in the promoter and first exon regions of genes. Preliminary screening tests are crucial for informing patients whether they should pursue more involved testing. We selected RASSF1, previously demonstrated to be aberrantly methylated in liquid biopsies from breast cancer patients, as our gene of interest. Using CoBRA as our method for methylation quantification, we designed unique primer sets that amplify a portion of the CpG island spanning the 5′ end of the RASSF1 first exon. We integrated the CoBRA approach with a microfluidics-based electrophoresis quantification system (LabChip) and optimized the assay such that insightful results could be obtained without post-PCR purification or concentration, two steps traditionally included in CoBRA assays. Circumventing these steps resulted in a decreased turnaround time and mitigated the laboratory machinery and reagent requirements. Our streamlined technique has an estimated limit of detection of 9.1 ng/μL of input DNA and was able to quantify methylation with an average error of 4.3%. Full article
(This article belongs to the Section Medical Biotechnology)
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14 pages, 1653 KiB  
Article
Novel CaLB-like Lipase Found Using ProspectBIO, a Software for Genome-Based Bioprospection
by Gabriela C. Brêda, Priscila E. Faria, Yuri S. Rodrigues, Priscila B. Pinheiro, Maria Clara R. Nucci, Pau Ferrer, Denise M. G. Freire, Rodrigo V. Almeida and Rafael D. Mesquita
BioTech 2023, 12(1), 6; https://doi.org/10.3390/biotech12010006 - 06 Jan 2023
Viewed by 2276
Abstract
Enzymes have been highly demanded in diverse applications such as in the food, pharmaceutical, and industrial fuel sectors. Thus, in silico bioprospecting emerges as an efficient strategy for discovering new enzyme candidates. A new program called ProspectBIO was developed for this purpose as [...] Read more.
Enzymes have been highly demanded in diverse applications such as in the food, pharmaceutical, and industrial fuel sectors. Thus, in silico bioprospecting emerges as an efficient strategy for discovering new enzyme candidates. A new program called ProspectBIO was developed for this purpose as it can find non-annotated sequences by searching for homologs of a model enzyme directly in genomes. Here we describe the ProspectBIO software methodology and the experimental validation by prospecting for novel lipases by sequence homology to Candida antarctica lipase B (CaLB) and conserved motifs. As expected, we observed that the new bioprospecting software could find more sequences (1672) than a conventional similarity-based search in a protein database (733). Additionally, the absence of patent protection was introduced as a criterion resulting in the final selection of a putative lipase-encoding gene from Ustilago hordei (UhL). Expression of UhL in Pichia pastoris resulted in the production of an enzyme with activity towards a tributyrin substrate. The recombinant enzyme activity levels were 4-fold improved when lowering the temperature and increasing methanol concentrations during the induction phase in shake-flask cultures. Protein sequence alignment and structural modeling showed that the recombinant enzyme has high similarity and capability of adjustment to the structure of CaLB. However, amino acid substitutions identified in the active pocket entrance may be responsible for the differences in the substrate specificities of the two enzymes. Thus, the ProspectBIO software allowed the finding of a new promising lipase for biotechnological application without the need for laborious and expensive conventional bioprospecting experimental steps. Full article
(This article belongs to the Special Issue Bioinformatics: Present and Future Biotechnology)
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11 pages, 2319 KiB  
Article
Effects of Pseudomonas sp. OBA 2.4.1 on Growth and Tolerance to Cadmium Stress in Pisum sativum L.
by Liliya Khakimova, Olga Chubukova, Zilya Vershinina and Dilara Maslennikova
BioTech 2023, 12(1), 5; https://doi.org/10.3390/biotech12010005 - 03 Jan 2023
Cited by 2 | Viewed by 2097
Abstract
Cadmium stress is a barrier to crop production, yield, quality, and sustainable agriculture. In the current study, we investigated the characteristics of bacterial strain Pseudomonas sp. OBA 2.4.1 under cadmium (CdCl2) stress and its influence on Cd stresses in pea ( [...] Read more.
Cadmium stress is a barrier to crop production, yield, quality, and sustainable agriculture. In the current study, we investigated the characteristics of bacterial strain Pseudomonas sp. OBA 2.4.1 under cadmium (CdCl2) stress and its influence on Cd stresses in pea (Pisum sativum L.) seedlings. It was revealed that strain OBA 2.4.1 is tolerant of up to 2 mM CdCl2, and seed treatment with the bacterium enhanced pea plant growth (length of seedlings) under 0.5 mM cadmium stress. This bacterial strain showed plant growth-promoting properties, including biofilm formation and siderophore activity. An important advantage of the studied strain OBA 2.4.1 is its ability to colonize the plant roots. Moreover, the inoculation with strain OBA 2.4.1 significantly reduced oxidative stress markers in pea seedlings under cadmium stress. These findings suggest that cadmium stress-tolerant strain OBA 2.4.1 could enhance pea plant growth by mitigating stress-caused damage, possibly providing a baseline and eco-friendly approach to address heavy metal stress for sustainable agriculture. Full article
(This article belongs to the Section Environmental Biotechnology)
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21 pages, 3508 KiB  
Review
Herbs as a Source for the Treatment of Polycystic Ovarian Syndrome: A Systematic Review
by Jada Naga Lakshmi, Ankem Narendra Babu, S. S. Mani Kiran, Lakshmi Prasanthi Nori, Nageeb Hassan, Akram Ashames, Richie R. Bhandare and Afzal B. Shaik
BioTech 2023, 12(1), 4; https://doi.org/10.3390/biotech12010004 - 03 Jan 2023
Cited by 7 | Viewed by 10955
Abstract
Background: Polycystic ovarian syndrome (PCOS) is a neuroendocrine metabolic disorder characterized by an irregular menstrual cycle. Treatment for PCOS using synthetic drugs is effective. However, PCOS patients are attracted towards natural remedies due to the effective therapeutic outcomes with natural drugs and the [...] Read more.
Background: Polycystic ovarian syndrome (PCOS) is a neuroendocrine metabolic disorder characterized by an irregular menstrual cycle. Treatment for PCOS using synthetic drugs is effective. However, PCOS patients are attracted towards natural remedies due to the effective therapeutic outcomes with natural drugs and the limitations of allopathic medicines. In view of the significance of herbal remedies, herein, we discuss the role of different herbs in PCOS. Methods: By referring to the Scopus, PubMed, Google Scholar, Crossref and Hinari databases, a thorough literature search was conducted and data mining was performed pertaining to the effectiveness of herbal remedies against PCOS. Results: In this review, we discuss the significance of herbal remedies in the treatment of PCOS, and the chemical composition, mechanism of action and therapeutic application of selected herbal drugs against PCOS. Conclusions: The present review will be an excellent resource for researchers working on understanding the role of herbal medicine in PCOS. Full article
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14 pages, 1047 KiB  
Article
Molecular and Physiological Variability in Bread Wheat and Its Wild Relative (Aegilops tauschii Coss.) Species under Water-Deficit Stress Conditions
by Zahra Khodadadi, Mansoor Omidi, Alireza Etminan, Asa Ebrahimi and Alireza Pour-Aboughadareh
BioTech 2023, 12(1), 3; https://doi.org/10.3390/biotech12010003 - 28 Dec 2022
Cited by 1 | Viewed by 1888
Abstract
Aegilops and Triticum spp. are two ideal gene pools for the breeding purposes of wheat. In this study, a set of Iranian accessions of Aegilops tauschii Coss. and Triticum aestivum L. species were evaluated in terms of some physiological and biochemical features under [...] Read more.
Aegilops and Triticum spp. are two ideal gene pools for the breeding purposes of wheat. In this study, a set of Iranian accessions of Aegilops tauschii Coss. and Triticum aestivum L. species were evaluated in terms of some physiological and biochemical features under control and water-deficit stress conditions. Moreover, several simple sequence repeat (SSR) markers were employed to identify marker loci associated with the measured traits. The results indicated that water-deficit stress significantly affected all measured traits and the highest reductions due to water-deficit were recorded for shoot fresh and dry biomasses (SFB and SDB), stomatal conductance (Gs), leaf relative water content (RWC), and chlorophyll b content (Chl b). In molecular analysis, 25 SSR markers generated 50 fragments, out of which 49 fragments (98%) were polymorphic. Furthermore, the genetic variation observed within species is more than between species. The results of cluster and Bayesian model analysis classified all evaluated accessions into three main clusters. Under control and water-deficit stress conditions, 28 and 27 significant marker-trait associations (MTAs) were identified, respectively. Furthermore, 10 MTAs showed sufficiently stable expression across both growth conditions. Of these, the markers Xgwm-111, Xgwm-44, Xgwm-455, Xgwm-272, and Xgwm-292 were associated with multiple traits. Hence, these markers could serve as useful molecular tools for population characterization, gene tagging, and other molecular breeding studies. Full article
(This article belongs to the Special Issue New Technologies in Agricultural and Plant Biotechnology)
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12 pages, 3193 KiB  
Article
Development of Targeted Protein-Displaying Technology with a Novel Carbon Material
by Akihito Nakanishi, Naotaka Yamamoto, Yuri Sakihama, Tomoya Okino and Naoki Matoba
BioTech 2023, 12(1), 2; https://doi.org/10.3390/biotech12010002 - 25 Dec 2022
Viewed by 2019
Abstract
This study reports a new carbon material and its specific display of targeted protein. The properties of the carbon materials fabricated with carbon black MOGUL® were analyzed. The carbon materials were spherical structures with 55.421 µm as a median value. The specific [...] Read more.
This study reports a new carbon material and its specific display of targeted protein. The properties of the carbon materials fabricated with carbon black MOGUL® were analyzed. The carbon materials were spherical structures with 55.421 µm as a median value. The specific surface area, pore volume, average pore diameter, and total of the acidic functional group were 130 m2·g−1, 0.55 cm3·g−1, 17.2 nm, and 0.29 mEq·g−1, respectively. The adsorption–desorption isoform of the carbon materials showed type IV of the hysteresis loop as defined by IUPAC, indicating non-uniform mesoporous structures (2–50 nm). The distribution of the log differential pore volume also indicated non-uniform porous structures because (i) the difference between the average pore size and the most frequent pore size was significant and (ii) the σ value was larger than the average value regarding the pore sizes. However, 10–90% of the integrated values of the log differential pore volume were 57.4% of the total integrated values, and the distribution was similar to the Gauss distribution model. Although the value of the total of the acidic functional group was 2.5–5.4 times lower than the values of the HPLC columns, the carbon materials require good scaffold quality rather than good HPLC quality. Therefore, the amounts could be enough for the scaffold of biotin hydrazide. To demonstrate the property of displaying the targeted proteins, carbon materials displaying biotin hydrazide by covalent bonding were prepared and avidin-labeled horse radish peroxidase (HRP) was bound to the biotin region. The carbon materials were porous structures, so the unspecific adsorption of HRP was estimated. Then, the maintenance ratios of HRP activities were analyzed in the repeated-use-with-wash processes after each evaluation, resulting in the activities of HRP on the carbon materials being treated with biotin hydrazide being significantly maintained compared to that of the ones without biotin hydrazide. The study revealed the properties of the carbon materials and indicated the display of HRP, suggesting that the carbon materials could be a new material for displaying targeted proteins. Full article
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9 pages, 210 KiB  
Article
Genome Editing in Human Gametes and Embryos: The Legal Dimension in Europe
by Takis Vidalis
BioTech 2023, 12(1), 1; https://doi.org/10.3390/biotech12010001 - 23 Dec 2022
Cited by 7 | Viewed by 2153
Abstract
To date, the legal aspects of the ongoing debate on the application of genome editing in human gametes and early embryos have attracted little attention. In Europe, this seems to have changed with a recent official position that clarifies the meaning of the [...] Read more.
To date, the legal aspects of the ongoing debate on the application of genome editing in human gametes and early embryos have attracted little attention. In Europe, this seems to have changed with a recent official position that clarifies the meaning of the relevant provision of the common legal instrument on Human Rights and Biomedicine (Oviedo Convention). This provision explicitly prohibits modifications to the genome of future persons and adoptes a precautionary stance with regard to genetic interventions in the human reproductive material. In this article, we examine relevant interpretative options, following the new official clarifications, focusing on the research/clinical application distinctions that characterize their approach. From this viewpoint, we propose an approach that favors basic research activities involving genome editing, even for exploring potential clinical applications under conditions of safety, which may justify a future legislative amendment. Furthermore, we explore the patenting issue, based on the current approach of European case law, and give reasons that may justify patent rights in this ethically sensitive area. Full article
(This article belongs to the Section Biotechnology Regulation)
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