미생물 셀룰로오스 생산을 위한 당밀의 전처리 및 생산된 셀룰로오스의 물리화학적 특성

Utilization of acetate buffer to improve bacterial cellulose production by Gluconacetobacter xylinus

Metabolomic profiling coupled with metabolic network reveals differences in Gluconacetobacter xylinus from static and agitated cultures

This study aimed to isolate an efficient bacterial cellulose (BC)-producing strain from rotten mango, optimize BC production though single-factor tests followed by Box-Behnken design (BBD) under agitated culture conditions using tobacco waste extract as the medium, and conduct the characterization of BC via Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), and X-ray diffraction (XRD). Komagataeibacter sp. XMZ1 with a high-efficiency BC biosynthesis capacity was successfully isolated from rotten mango. Subsequently, BBD was utilized to optimize the co-supplementation of lactic acid, ethanol, and yeast extract, demonstrating this statistical approach to be a reliable tool for both optimization and predictive modeling of BC yield. The optimized medium containing 2.1g/L lactic acid, 1% (v/v) ethanol, and 2g/L yeast extract resulted in a 2.1-fold enhancement of BC production, increasing the yield from 4.20g/L to 8.85g/L. The characterization of BC through FTIR, SEM and XRD revealed that BC produced under agitated culture conditions maintained similar chemical functional groups as the BC produced under static culture. However, the agitated culture products exhibited a more porous fibrillar network and a reduced crystallinity index. This present study provides valuable insights into BC production under agitated culture conditions by making use of tobacco waste. Additionally, it offers BC structural characterizations that are relevant to potential industrial applications.

Some bacterial strains such as Komagataeibacter xylinus are able to produce cellulose as an extracellular matrix. In comparison to wood-based cellulose, bacterial cellulose (BC) holds interesting properties such as biodegradability, high purity, water-holding capacity, and superior mechanical and structural properties. Aiming toward improvement in BC production titer and tailored alterations to the BC film, we engineered K. xylinus to overexpress partial and complete bacterial cellulose synthase operon that encodes activities for BC production. The changes in cell growth, end metabolite, and BC production titers from the engineered strains were compared with the wild-type K. xylinus. Although there were no significant differences between the growth of wild-type and engineered strains, the engineered K. xylinus strains demonstrated faster BC production, generating 2-4-fold higher production titer (the highest observed titer was obtained with K. xylinus-bcsABCD strain producing 4.3 ± 0.46 g/L BC in 4 days). The mechanical and structural characteristics of cellulose produced from the wild-type and engineered K. xylinus strains were analyzed with a stylus profilometer, in-house built tensile strength measurement system, a scanning electron microscope, and an X-ray diffractometer. Results from the profilometer indicated that the engineered K. xylinus strains produced thicker BC films (wild type, 5.1 μm, and engineered K. xylinus strains, 6.2-10.2 μm). Scanning electron microscope revealed no principal differences in the structure of the different type BC films. The crystallinity index of all films was high (from 88.6 to 97.5%). All BC films showed significant piezoelectric response (5.0-20 pC/N), indicating BC as a promising sensor material.

Bacterial Cellulose (BC) offers a wide range of applications across various industries, including food, biomedical, and textiles, owing to its distinctive properties. Its unique 3D reticulated network of cellulose nanofibers, imparts excellent mechanical qualities, a high water-holding capacity, and thermal stability. Additionally, it possesses remarkable biocompatibility, biodegradability, high crystallinity, and purity. These attributes have offered significant interest in BC within both academic and industrial sectors. However, BC production is associated with high costs due to the use of expensive growth media and low yields. The study reports the potential of our indigenous isolate, Komagataeibacter saccharivorans BC-G1, as BC producer. Statistical optimization of BC production was carried out using Placket-Burman design and Central composite design, by selecting different parameters. Eight significant factors such as temperature, pH, glucose, yeast, peptone, acetic acid, incubation time and % inoculum were studies using ANOVA-based response surface methodology. Results showed that BC yield (8.5g/L) with 1.8-fold after optimization of parameters. Maximum cellulose production (8.5 ± 1.8g/L) was obtained using 2% glucose, 0.3% yeast extract, 0.3% peptone, 0.75% (v/v) acetic acid at pH 7.0 for 10 days of incubation with 4% inoculum at 25°C under static culture. Main effect graph showed incubation time and acetic acid concentration as the most significant parameters affecting BC production in our study. The physicochemical characterization of produced BC was done using FTIR, XRD and SEM techniques.

Bacterial cellulose (BC) is a potential eco-friendly biopolymer. BC has higher crystallinity and purity compared to plant cellulose. Scientific studies on the production of BC from lactic acid bacteria (LAB) are minimal compared to other common bacteria such as Acetobacter xylinum. LAB was screened and isolated from different tissues of Mimosa pudica (medicinal plant) using MRS broth and agar as the selective medium. LAB isolates were subjected to 16S rRNA gene sequencing of all the bacterial isolates. BC was produced from all LAB isolates by incubating at 30 °C for 14 days in herbal tea medium (Strobilanthes crispus) and HS medium (control) with 130 r.p.m agitation. BC produced by two selected bacterial isolates was characterized using FESEM, FTIR, XRD, and TGA. Molecular analysis of the 16S rRNA gene of all the potential LAB isolates shows 99.86 - 100% identity to 16S rRNA sequences of other Lactobacillus plantarum strains. Two selected L. plantarum strains (LBM001 & LBM004) produce BC in sphere-like particles with a 1.4 to 2.2 µm diameter range of microfiber. FTIR analysis shows that BC produced by LBM001 and LBM004 have four similar cellulose regions identified in cellulose from other sources, which are O-H stretch (3400-330 cm-1), C-H stretch (2970-2800 cm-1), O-H bending (1620cm-1) and C-O-C stretch (1100-1073 cm-1). XRD analysis shows BC produced by the L. plantarum strains consists of two different XRD peaks at the 2θ angle of 21.53° and 21.85° instead of a single peak (22.76°) identified in the BC produced by A. xylinum and plant cellulose. A similar TG and DTG curved pattern was detected in the BC produced by the L. plantarum strains with the BC produced by A. xylinum and plant cellulose. The LAB isolates from M. pudica have potential in BC production based on the multiple characterization studies.

The objective of the work is to examine the potential utilization of Palmyra palm jaggery (PPJ) for the enhancement of bacterial cellulose (BC) production byGluconacetobacter liquefaciens. To evaluate the culturing condition, the production of BC fermentation was carried out in batch mode using different carbon sources namely glucose, sucrose and PPJ. PPJ in the HS medium (PHS medium) resulted maximum concentration of BC (14.35 ± 0.18g/L) under shaking condition than other carbon sources in HS medium. The influence of different medium variables including initial pH and nitrogen sources on BC production was investigated using PHS medium under shaking condition. The maximum BC concentration of 17.79 ± 2.4g/L was obtained in shaking condition at an initial pH of 5.6 using yeast extract as nitrogen source. Stoichiometric equation for the cell growth and BC synthesis was developed using elemental balance approach. The metabolic heat of reaction (40kcal generated per liter of medium) was evaluated using electron balance approach. Based on the process economic analysis and the yield of BC during the fermentation, PHS medium without nitrogen source could be a promising cost-effective nutrient than HS medium. Thermal stability, crystallinity index and structural characterizations of produced BC using PPJ medium were evaluated using TGA, XRD and FTIR and the obtained results were compared with HS medium containing glucose and sucrose.

Biofilm reactors show several advantages over suspended cell reactors, especially in their biomass density, operation convenience, and production yield. The structure of biofilm matrix contributes to high resistance of microorganisms to extreme conditions of pH, temperature, toxic chemicals, and mechanical shocks. Submerged fermentation has been used for BC production due to its feasibility of large-scale production in industry. The purpose of this research topic is to improve the rate of bacterial cellulose (BC) production by exploring new culture methods with a biofilm reactor. Biofilm reactor included a plastic composite support (PCS) which was attached on agitation shaft to promote effective nutrient delivery and to control biofilm thickness. BC harvested from PCS biofilm reactor was evaluated in terms of the physical and mechanical properties, including the degree of crystallinity and crystal size by x-ray diffraction (XRD), the water content and degradation characteristics using thermogravimetric analysis (TGA), nanostructure using field emission scanning electron microscopy (FESEM), and tensile strength using a dynamic mechanical analyzer (DMA). Among thirteen types of PCS, PCS blended with dried soybean hulls, defatted soybean flour, yeast extract, dried bovine red blood cells, and mineral salts (SFYR+) yielded the highest BC (7.05 g/l) which is 2.5 fold compared to control (2.82 g/l). The XRD studies indicated that crystallinity of BC pellicle is about 90%. FESEM image showed Acetobacter xylinum grew and produced BC on the PCS. TGA results showed that BC pellicle on the PCS had 95% water content. The results clearly demonstrated that implemented with PCS in agitated bioreactor can enhance BC production.

To offset the negative effects of aeration on bacterial cellulose (BC) production by acetic acid bacteria using enmeshed cellulose microfibrils (CM) on luffa sponge matrices (LSM). The CM were enmeshed on LSM (LSM-CM). The optimal amount of LSM-CM was determined for BC production under aerated conditions. Without LSM-CM, no BC was produced in seven out of nine production cycles at the highest aeration rate (9lmin-1 ). However, with 0·5% LSM-CM and an aeration rate of 3lmin-1 , a satisfactory oxygen transfer coefficient was achieved, and also a good yield of BC (5·24gl-1 ). Moreover, the LSM-CM was able to be recycled through nine consecutive BC production cycles. The highest BC yields (from 5·8±0·4 to 6·6±0·4gl-1 ) were associated with high bacterial biomass and this was confirmed by scanning electron microscopy. We confirm that LSM-CM works well as a starter. Microenvironments low in dissolved oxygen within the matrices of LSM-CM are important for BC production under aeration conditions. The LSM-CM provides a microenvironment which offsets the negative effects of aeration on BC production. A sustainable, economic process for mass BC production is described using recycled LSM-CM with aeration.

The advances in bioprocess design, genetic engineering, and media optimization have enabled enhanced bacterial cellulose (BC) production and its application for diverse purposes. Following the requirements of a bioeconomy, numerous approaches were conducted to investigate alternative carbon or nitrogen sources from industrial by-products for BC biosynthesis. They can, however, not only affect the BC production but also its properties. Beet molasses, vinasse, and waste beer fermentation broth (WBFB) have thus been investigated in single and combined approaches for their BC production potential and effects on structural properties using Komagataeibacter xylinus DSM 2325. Therefore, the composition of each complex component was initially analyzed for total organic carbon (TOC), total bound nitrogen (TNb), sugars, organic acids, and alcohols. The polymer properties were characterized via gel permeation chromatography and X-ray diffraction. In dynamic shake flask cultivations, the exchange of Hestrin-Schramm (HS) medium components for a combination of all three complex substrates on a TOC- or TNb-based quantity resulted in the highest BC concentration (8.2 g L−1). Comparable concentrations were achieved when combining molasses and WBFB (8.1 g L−1). Each investigated complex component led to differing degrees of polymerization (DPn: 2751−4601) and BC crystallinities (26−58%) in comparison to HS medium. Beet molasses and vinasse were found to decrease the polymer crystallinity but induce higher DPn whereas the opposite occurred for WBFB. This study thus highlights beneficial effects of food industry by-products for BC biosynthesis and elucidates concomitantly occurring structural polymer alterations to enable further studies dealing with alternative substrates for structurally tailored BC production.

Antarctic microorganisms have developed extraordinary strategies for adaptation. They have also demonstrated the ability to produce various biopolymers in response to environmental stress. The demand for biopolymers is constantly increasing and is expected to grow further. Among emerging biomaterials, bacterial cellulose (BC) is generating significant interest due to its unique characteristics that distinguish it from plant-based cellulose. BC exhibits higher purity, water-holding capacity, and tensile strength compared to its plant-based counterpart. Furthermore, BC can be obtained through environmentally friendly protocols. Several bacterial strains have already been identified as cellulose producers, including Komagataeibacter xylinus. In this study, a marine bacterial strain named Pseudomonas sp. ef1, isolated from a consortium associated with the Antarctic ciliate Euplotes focardii, was tested for cellulose production. We found that this Antarctic Pseudomonas can produce BC in conditions that appear unique to this bacterial strain. Furthermore, the final BC product is structurally different from that obtained from the well-known BC producer Komagataeibacter xylinus. Additionally, a putative cellulose synthase was identified from the Pseudomonas sp. ef1 genome, exhibiting unique characteristics that may account for the unique BC production capability of this Antarctic marine Pseudomonas. The versatility of BC opens numerous applications, including in papermaking, food, pharmaceutical, and biomedical sectors.

10 - Production of bio-cellulose from renewable resources: Properties and applications

Bacterial cellulose (BC) has been extensively utilized in a wide range of applications specifically in the biomedical field thanks to its excellent physico-chemical and biological features. The major limitation restricting its application in certain areas is its high production cost. Its widespread applications demand exploration of alternative production media compared to the existing expensive ones. Herein, an effort has been made to utilize waste and cheaply available local resources including; waste (expired) orange juice (WOJ), sugarcane juice (SC) and coconut water (CW) as alternative media for BC production in comparison to the synthetic media (control). Waste and cheap resources were collected from the local market, screened filtered and optimized for the development of BC culture media. BC production from all media was observed under static cultivation for 10 days. The results indicated 2.75, 2.56, 3.32 and 1.68 g/L BC production that corresponded to 27.5%, 21.7 %, 20.1 % and 31.6 % sugar to BC conversion from control, WOJ, SC and CW media, respectively. Morphology and crystalline features of produced BC samples were observed through FE-SEM and XRD analysis. It was noteworthy that BC produced from all alternative sources indicated high water holding capabilities (WHC) and water retention time (WRT) that augment their applicability in drug delivery and wound healing applications. The BC production from cheap resources and its high physical, mechanical and biological properties can be of high interest for scaling up and commercialization of BC production processes. Furthermore, its liquidabsorbing capabilities and retention time can help in drug carrying and medical application.

Papaya peel is a fruit waste that was usually dumped into the environment. One way to utilize papaya peel waste was to convert it into a fermented product like bacterial cellulose. In this study, the effect of various bacterial cellulose nitrogen sources such as bean sprouts, coconut milk and urea food grade were used to produce bacterial cellulose from papaya peel. In addition, the determination of the bacterial cellulose kinetic parameters and techno-economic analysis were also evaluated. The results from this research showed that urea food grade as nitrogen source produced highest bacterial cellulose. Longer fermentation time produced higher bacterial cellulose and lower water content in bacterial cellulose production. From kinetic model optimization with bacterial cellulose data, kinetic parameters such as maximum specific growth rate (µmax), monod constant (Ks), cell death rate constant (Kd), and cell maintenance constant (m) were 0.06 (day-1), 1.25 (g/L), 0.117 (day-1), and 0.568 (day-1). Techno-economic evaluation showed that bacterial cellulose production with recycle medium stage produced high profitability. Profitability parameters value such as return of investment (ROI), payback period (PBP), net present value (NPV), and internal rate of return (IRR) were 75.92%, 1.01 years, US$ 1,839,257,209, and 76.94%. This research showed that higher bacterial cellulose yield can be produced from papaya peel waste and urea food grade. Techno-economic simulations showed that large-scale production of bacterial cellulose from papaya peel waste can be profitable by recycle the fermentation medium.

Effect of co-culture of Komagataeibacter nataicola and selected Lactobacillus fermentum on the production and characterization of bacterial cellulose