Carbohydrate metabolite pathways and antibiotic production variations of a novel Streptomyces sp. M3004 depending on the concentrations of carbon sources

To determine the variations of growth, some key enzyme activities such as glucose kinase (GK), glucose-6-phosphate dehydrogenase (G6PDH), α-ketoglutarate dehydrogenase (KGDH), and isocitrate lyase (ICL) besides metabolite levels of pyruvate and antibiotic production of newly isolated Streptomyces sp. M3004 were grown in culture media which contain 10–20 g/l concentration with either glucose or glycerol as carbon source. Biomass and intracellular glucose and glycerol levels of Streptomyces sp. M3004 showed positive correlation with the concentration of these carbon sources, and these levels were higher in glucose compared with the glycerol-supplemented mediums. GK, G6PDH, and KGDH activities showed marked correlation with the concentration of both glucose and glycerol, and the activity levels were 4.14-, 1.47-, and 1.27-fold higher in glucose than glycerol. A key enzyme of the glyoxalate cycle, ICL activities decreased with increasing glucose concentrations from 10 to 20 g/l, but increased up to 15 g/l of glycerol. The positive correlations were also determined between intracellular glucose and glycerol levels besides pyruvate and protein variations with respect to concentrations of the carbon sources. Antibacterial activities of Streptomyces sp. M3004 reached maximum on the stationary phase, while it did not change significantly with respect to glucose and glycerol.     

Contribution of Glycerol on Production of Poly(γ-Glutamic Acid) in <Emphasis Type="Italic">Bacillus subtilis</Emphasis> NX-2

Glycerol would stimulate the production of poly(γ-glutamic acid) (γ-PGA) and decrease its molecular weight in Bacillus subtilis NX-2. When 20 g/l glycerol was added in the medium, the yield of γ-PGA increased from 26.7 ± 1.0 to 31.7 ± 1.3 g/l, and molecular weight of γ-PGA decreased from 2.43 ± 0.07 × 10⁶ to 1.86 ± 0.06 × 10⁶ Da. In addition, it was found that the decrease of γ-PGA chain length by glycerol would lead to the decrease of broth viscosity during the fermentation and enhanced the uptake of substrates, which could not only improve cell growth but also stimulate γ-PGA production. Moreover, it was also found that glycerol could effectively regulate molecular weight between 2.43 ± 0.07 × 10⁶ and 1.42 ± 0.05 × 10⁶ Da with the concentration ranging from 0 to 60 g/l. This was the first time to discover such contribution of glycerol on γ-PGA production in Bacillus genus. And the effects of glycerol on molecular weight of γ-PGA would be developed to be an approach for the regulation of microbial γ-PGA chain length, which is of practical importance for future commercial development of this polymer.     

Enhancement of Human γ-Interferon Production in Recombinant <Emphasis Type="Italic">E. coli</Emphasis> Using Batch Cultivation

Development of inexpensive and simple culture media and appropriate induction conditions are always favorable for industry. In this research, chemical composition and stoichiometric data for γ-interferon production and recombinant Escherichia coli growth were used in order to achieve a simple medium and favorable induction conditions. To achieve this goal, the effects of medium composition and induction conditions on the production of γ-interferon were investigated in batch culture of E. coli BL21 (DE3) [pET3a-ifnγ]. These conditions were considered as suitable conditions for the production of γ-interferon: 2.5× M9 medium, supplemented with a mixture of amino acids (milligram per liter), including glutamic acid 215, aspartic acid 250, lysine 160, and phenylalanine 90, and induction at late-log phase (OD₆₀₀ = 4.5). Under these conditions, dry cell weight of 6 ± 0.2 g/l and γ-interferon concentration of 2.15 ± 0.1 g/l were obtained. Later, without changing the concentration ratio of amino acids and glucose, the effect of increase in the primary glucose concentration on productivity of γ-interferon was investigated. It was found that 25 g/l glucose will result in maximum attainable biomass and recombinant human γ-interferon. At improved conditions, a dry cell weight of 14 ± 0.2 g/l, concentration and overall productivity of γ-interferon 4.2 ± 0.1 g/l and 420 ± 10 mg/l h, respectively, were obtained.     

Production and Characterization of an Alkaline Thermostable Crude Lipase from an Isolated Strain of <Emphasis Type="Italic">Bacillus cereus</Emphasis> C<Subscript>7</Subscript>

A bacterial strain isolated from spoiled coconut and identified as Bacillus cereus was found capable of producing alkaline thermostable extracellular lipase. Optimum temperature, time, and pH for enzyme substrate reaction were found to be 60 °C, 10 min, and 8.0 respectively. Common surfactants except Triton X 100 and cetyltrimethylammonium bromide have no or very little inhibitory effects on enzyme activity. The enzyme was found to be stable in presence of oxidizing agents and protease enzyme. The maximum lipase production was achieved at 30–33 °C, pH 8.0 on 24 h of fermentation using 50 ml medium in a 250-ml Erlenmeyer flask. The superior carbon and nitrogen sources for lipase production were starch (2%) and ammonium sulfate (nitrogen level 21.2 mg/100 ml), peptone (nitrogen level 297 mg/100 ml), and urea (nitrogen level 46.62 mg/100 ml) in combination, respectively. The maximum enzyme activity obtained was 33 ± 0.567 IU/ml.     

Banana Peel: A Potential Substrate for Laccase Production by <Emphasis Type="Italic">Aspergillus fumigatus</Emphasis> VkJ2.4.5 in Solid-State Fermentation

In solid-state fermentation, among various solid supports evaluated, banana peel was found to be an ideal support and resulted into higher levels of laccase (6281.4 ± 63.60 U l⁻¹) along with notable levels of manganese peroxidase production (1339.0 ± 131.23 U l⁻¹) by Aspergillus fumigatus VkJ2.4.5. Maximum levels of laccase was achieved under derived conditions consisting of 80% of moisture level, 6 days of incubation period, 6% inoculum level, and an aeration level of 2.5 l min⁻¹. A column-tray bioreactor was designed to scale up and economize the enzyme production in three successive cycles of fermentation using the same fungal biomass. Thermal and pH stability profiles revealed that enzyme was stable up to 50°C and at varying pH range from 5–9 for up to 2 h. The apparent molecular weight of laccase was found to be 34 ± 1 kDa. MALDI-TOF/TOF analysis of the protein showed significant homology with maximum identity of 67% to other laccases reported in database.     

Stimulated Production of Triterpenoids of <Emphasis Type="Italic">Ganoderma lucidum</Emphasis> by an Ether Extract from the Medicinal Insect, <Emphasis Type="Italic">Catharsius molossus</Emphasis>, and Identification of the Key Stimulating Active Components

The medicinal fungus Ganoderma lucidum was inoculated into the media with and without supplementation of medicinal insect extracts to screen stimulators from Chinese medicinal insects for mycelial growth and triterpenoids production in submerged fermentation. The methanol and ether extracts of the tested insects had no significant stimulatory effect on the mycelial biomass production (P > 0.05), and those of H. remigator and Mylabris phalerata markedly inhibited the mycelial growth. However, the ether extract of Catharsius molossus at a concentration of 200 mg l⁻¹ led to a significant increase in triterpenoids concentration from 231.7 ± 9.77 to 313.7 ± 10.6 mg l⁻¹ (P < 0.01). Analysis of fermentation kinetics of G. lucidum suggests that glucose concentration in the extract of C. molossus-added group decreased more quickly as compared to the control group from day 2 to day 7 of fermentation process, while the triterpenoids biosynthesis was promoted at the same culture period. However, the culture pH profile was not affected by the addition of the extract. Chemical study of the extract show that cis-9,10-methylenehexadecanoic acid (9,10-MEA) and hexadecanoic acid (especially 9,10-MEA) were the key active compounds of the extract responsible for the stimulatory effect on the triterpenoids production.     

Xylanase Production by <Emphasis Type="Italic">Penicillium canescens</Emphasis> on Soya Oil Cake in Solid-State Fermentation

There is an increasing interest for the organic residues from various sectors of agriculture and industries over the past few decades. Their application in the field of fermentation technology has resulted in the production of bulk chemicals and value-added products such as amino acid, enzymes, mushroom, organic acids, single-cell protein, biologically active secondary metabolites, etc. (Ramachandran et al., Bioresource Technology 98:2000–2009, 2007). In this work, the production of extracellular xylanase by the fungus Penicillium canescens was investigated in solid-state fermentation using five agro-industrial substrates (soya oil cake, soya meal, wheat bran, whole wheat bran, and pulp beet). The best substrate was the soya oil cake. In order to optimize the production, the most effective cultivation conditions were investigated in Erlenmeyer flasks and in plastic bags with 5 and 100 g of soya oil cake, respectively. The initial moisture content, initial pH, and temperature of the culture affected the xylanase synthesis. The optimal fermentation medium was composed by soya oil cake crushed to 5 mm supplemented with 3% and 4% (w/w) of casein peptone and Na₂HPO₄.2H₂O. After 7 days of incubation at 30 °C and under 80% of initial moisture, a xylanase production level of 18,895 ± 778 U/g (Erlenmeyer flasks) and 9,300 ± 589 U/g (plastic bags) was reached. The partially purified enzyme recovered by ammonium sulfate fractionation was completely stable at freezing and refrigeration temperatures up to 6 months and reasonably stable at room temperature for more than 3 months.     

Continuous production of succinic acid by a fumarate-reducing bacterium immobilized in a hollow-fiber bioreactor

Enterococcus faecalis RKY1, a fumarate-reducing bacterium, was immobilized in an asymmetric hollow-fiber bioreactor (HFBR) for the continuous production of succinic acid. The cells were inoculated into the shell side of the HFBR, which was operated in transverse mode. Since the pH values in the HFBR declined during continuous operation to about 5.7, it was necessary to change the feed pH from 7.0 to 8.0 after 24 h of operation in order to enhance production of succinic acid. During continuous operation with a medium containing fumarate and glycerol, the productivity of succinate was 3.0–10.9 g/(L·h) with an initial concentration of 30 g/L of fumarate, 4.9–14.9 g/(L·h) with 50 g/L of fumarate, and 7.2–17.1 g/(L·h) with 80 g/L of fumarate for dilution rates between 0.1 and 0.4 h⁻¹. The maximum productivity of succinate obtained by the HFBR (17.1 g of succinate /[L·h]) was 1.7 times higher than that of the batch bioconversions (9.9 g of succinate /[L·h]) with 80 g/L of fumarate. Furthermore, the long-term stability of the HFBR was demonstrated with a continuously efficient production of succinate for more than 15 d (360 h).     

The Effect of pH on Continuous Biohydrogen Production from Swine Wastewater Supplemented with Glucose

The effect of pH on hydrogen production from liquid swine manure supplemented with glucose by a mixed culture of fermentative bacteria in an anaerobic sequencing batch reactor was evaluated in this study. At 37 ± 1 °C, five pH values ranging from 4.7 to 5.9 at an increment of 0.3 were tested at a hydraulic retention time (HRT) of 16 h. The results showed that at this HRT, the optimal pH for hydrogen production was 5.0, under which the biogas comprised 33.57 ± 5.65% of hydrogen with a production rate of 8.88 ± 2.94 L-H₂/day and a yield of 1.48 ± 0.49 L-H₂/L liquid swine manure. The highest biomass concentration, highest butyric acid to acetic acid ratio, lowest propionic acid concentration, and the best stability were all found at pH 5.0, while the highest CH₄ productivity was found at pH 5.9. For efficient hydrogen production, oxygen content should be controlled under 2%, beyond which an inverse linear relationship (R ² = 0.986) was observed.     

Cellulases and Xylanases Production by <Emphasis Type="Italic">Penicillium echinulatum</Emphasis> Grown on Sugar Cane Bagasse in Solid-State Fermentation

To investigate the production of cellulases and xylanases from Penicillium echinulatum 9A02S1, solid-state fermentation (SSF) was performed by using different ratios of sugar cane bagasse (SCB) and wheat bran (WB). The greatest filter paper activity obtained was 45.82 ± 1.88 U gdm⁻¹ in a culture containing 6SCB/4WB on the third day. The greatest β-glucosidase activities were 40.13 ± 5.10 U gdm⁻¹ obtained on the third day for the 0SCB/10WB culture and 29.17 ± 1.06 U gdm⁻¹ for the 2SCB/8WB culture. For endoglucanase, the greatest activities were 290.47 ± 43.57 and 276.84 ± 15.47 U gdm⁻¹, for the culture 6SCB/4WB on the fourth and fifth days of cultivation, respectively. The greatest xylanase activities were found on the third day for the cultures 6SCB/4WB (36.38 ± 5.38 U gdm⁻¹) and 4SCB/6WB (37.87 ± 2.26 U gdm⁻¹). In conclusion, the results presented in this article showed that it was possible to obtain large amounts of cellulases and xylanases enzymes using low-cost substrates, such as SCB and WB.     

Ethanol Production from Cashew Apple Bagasse: Improvement of Enzymatic Hydrolysis by Microwave-Assisted Alkali Pretreatment

In this work, the potential of microwave-assisted alkali pretreatment in order to improve the rupture of the recalcitrant structures of the cashew able bagasse (CAB), lignocellulosic by-product in Brazil with no commercial value, is obtained from cashew apple process to juice production, was studied. First, biomass composition of CAB was determined, and the percentage of glucan and lignin was 20.54 ± 0.70% and 33.80 ± 1.30%, respectively. CAB content in terms of cellulose, hemicelluloses, and lignin, 19.21 ± 0.35%, 12.05 ± 0.37%, and 38.11 ± 0.08%, respectively, was also determined. Results showed that, after enzymatic hydrolysis, alkali concentration exerted influence on glucose formation, after pretreatment with 0.2 and 1.0 mo L⁻¹ of NaOH (372 ± 12 and 355 ± 37 mg g_glucan⁻¹) when 2% (w/v) of cashew apple bagasse pretreated by microwave-assisted alkali pretreatment (CAB-M) was used. On the other hand, pretreatment time (15–30 min) and microwave power (600–900 W) exerted no significant effect on hydrolysis. On enzymatic hydrolysis step, improvement on solid percentage (16% w/v) and enzyme load (30 FPU g_CAB-M⁻¹) increased glucose concentration to 15 g L⁻¹. The fermentation of the hydrolyzate by Saccharomyces cerevesiae resulted in ethanol concentration and productivity of 5.6 g L⁻¹ and 1.41 g L⁻¹ h⁻¹, respectively.     

Cloning,Expression, and Characterization of a Wide-pH-Range Stable Phosphite Dehydrogenase from <Emphasis Type="Italic">Pseudomonas</Emphasis> sp. K in <Emphasis Type="Italic">Escherichia coli</Emphasis>

A phosphite dehydrogenase gene (ptdhK) consisting of 1,011-bp nucleotides which encoding a peptide of 336 amino acid residues was cloned from Pseudomonas sp. K. gene ptdhK was expressed in Escherichia coli BL21 (DE3) and the corresponding recombinant enzyme was purified by metal affinity chromatography. The recombinant protein is a homodimer with a monomeric molecular mass of 37.2 kDa. The specific activity of PTDH-K was 3.49 U mg⁻¹ at 25 °C. The recombinant PTDH-K exhibited maximum activity at pH 3.0 and at 40 °C and displayed high stability within a wide range of pHs (5.0 to 10.5). PTDH-K had a high affinity to its natural substrates, with K _m values for sodium phosphite and NAD of 0.475 ± 0.073 and 0.022 ± 0.007 mM, respectively. The activity of PTDH-K was enhanced by Na⁺, NH₄⁺, Mg²⁺, Fe²⁺, Fe³⁺, Co²⁺, and EDTA, and PTDH-K exhibited different tolerance to various organic solvents.     

Determination of fatty acid compositions and cholesterol levels of some freshwater fish living in Porsuk Dam,Turkey

We determined the oil content, fatty acid composition, and cholesterol content of common carp (Cyprinus carpio), crucian carp (Carassius carassius), chub (Leusiscus cephalus), and tench (Tinca tinca) by GLC. The saturated fatty acids (SFA), monounsaturated fatty acids (MUFA), and polyunsaturated fatty acids (PUFA) levels were found to be 36.49%, 31.92%, 31.59% in common carp; 32.92%, 32.21%, and 34.87% in crucian carp; 36.19%, 32.91%, and 30.90% in chub; and 32.86%, 30.77%, and 36.37% in tench, respectively. The cholesterol (mg/100 g oil) levels of common carp, crucian carp, chub, and tench were determined by GLC methods as 119 ± 2.64 mg, 170.37 ± 2.36 mg, 94.68 ± 3.13 mg, and 179.84 ± 6.75 mg, respectively. Thus, the cholesterol contents of the analyzed freshwater fish species were low but their PUFA contents and nutritional values were high. Published in Khimiya Prirodnykh Soedinenii, No. 1, pp. 15–17, January–February, 2009.     

Preconditioning of Axillary Buds in Thidiazuron-Supplemented Liquid Media Improves In Vitro Shoot Multiplication in <Emphasis Type="Italic">Nyctanthes arbor-tristis</Emphasis> L.

An efficient tissue culture technology has been designed for mass multiplication of Nyctanthes arbor-tristis L. by preculturing nodal explants in thidiazuron (TDZ)-supplemented liquid Murashige and Skoog (MS) media. Direct inoculation of nodal segments on semi-solid MS medium augmented with various concentrations of TDZ (0.1 to 0.9 μM) produced shoots but with low regeneration response and few shoots per explant. Hence, nodal explants were pretreated with greater concentrations of TDZ (5 to 100 μM) in liquid MS media for different durations (4, 8, 12, and 16 days) with the aim of improving shoot regeneration response from cultured explants. After pretreatment, explants were transferred to agar-solidified hormone-free MS medium. Best response in terms of percent regeneration (94%), number of shoots per explant (20.00 ± 1.15), and greatest shoot length (7.23 ± 0.83 cm) were obtained with nodal segments pretreated in75 μM TDZ for 8 days. Similarly, root induction was obtained from pulse-treated microshoots for 24 h with 200 μM indole-3-butyric acid (IBA) followed by their transfer to 1/2 MS medium which produced an average of 5.50 ± 0.92 roots per microshoot. The rooted plantlets were transplanted to soil with 80% success rate.     

Studies on Silver Accumulation and Nanoparticle Synthesis By <Emphasis Type="Italic">Cochliobolus lunatus</Emphasis>

Development of reliable and eco-friendly processes for synthesis of metallic nanoparticles is an important step in the field of application of nanotechnology. Biological systems provide a useful option to achieve this objective. In this study, potent fungal strain was selectively isolated from soil samples on silver supplemented medium, followed by silver tolerance (100–1,000 ppm) test. The isolated fungus was subjected to morphological, 18S rRNA gene sequencing and phylogenic studies and confirmed as Cochliobolus lunatus. The silver accumulation and nanoparticle formation potential of wet cell mass of C. lunatus was investigated. The accumulation and nanoparticle formation by wet fungal cell mass with respect to pH change was also studied. The desorbing assay was used to recover accumulated silver from cell mass. C. lunatus was found to produce optimum biomass (0.94 g%) at 635 ppm of silver. Atomic absorption spectroscopy study showed that at optimum pH (6.5 ± 0.2), cell mass accumulates 55.6% of 100 ppm silver. SEM and FTIR studies revealed that the cell wall of C. lunatus is the site of silver sorption, and certain organic groups such as carbonyl, carboxyl, and secondary amines in the fungal cell wall have an important role in biosorption of silver in nanoform. XRD determined the FCC crystalline nature of silver nanoparticles. TEM analysis established the shape of the silver nanoparticles to be spherical with the presence of very small-sized nanoparticles. Average size of silver nanoparticles (14 nm) was confirmed by particle sizing system. This study reports the synthesis and accumulation of silver nanoparticles through reduction of Ag⁺ ions by the wet cell mass of fungus C. lunatus.     

Screening and Selection of Marine Isolate for l-Glutaminase Production and Media Optimization Using Response Surface Methodology

The current work details the screening of about 400 marine isolates from various marine niches, from which one isolate was finally selected based on the productivity of glutaminase (71.23 U/l). Further, biochemical identification tests and 16S rRNA sequencing identified this isolate to be Providencia sp. This isolate was taken up for further media optimization studies by using one-factor-at-a-time approach and subsequently by response surface methodology. A face centered central composite design was employed to investigate the interactive effects of four variables, viz., concentrations of glucose, methionine, urea, and succinic acid on glutaminase production. A significant influence of urea on glutaminase production was noted. Response surface methodology showed that a medium containing (g/l) glucose 10.0, urea 5.15, methionine 3.5, succinic acid 6.0, ammonium sulfate 2.5, and yeast extract 6.0 to be optimum for the production of glutaminase. The applied methodology was validated using this optimized media and enzyme activity 119 ± 0.12 U/l and specific activity of 0.63 U/mg protein after 28 h of incubation at 25 °C was obtained.     

A Superoxide Dismutase Purified from the Rhizome of <Emphasis Type="Italic">Curcuma aeruginosa</Emphasis> Roxb. as Inhibitor of Nitric Oxide Production in the Macrophage-like RAW 264.7 Cell Line

Superoxide dismutase (SOD, EC 1.15.1.1) is a metalloenzyme or antioxidant enzyme that catalyzes the disproportionation of the harmful superoxide anionic radical to hydrogen peroxide and molecular oxygen. Due to its antioxidative effects, SOD has long been applied in medicinal treatment, cosmetic, and other chemical industries. Fifteen Zingiberaceae plants were tested for SOD activity in their rhizome extracts. The crude homogenate and ammonium sulfate cut fraction of Curcuma aeruginosa were found to contain a significant level of SOD activity. The SOD enzyme was enriched 16.7-fold by sequential ammonium sulfate precipitation, diethylaminoethyl cellulose ion exchange, and Superdex 75 gel filtration column chromatography. An overall SOD yield of 2.51 % with a specific activity of 812.20 U/mg was obtained. The enriched SOD had an apparent MW of 31.5 kDa, as judged by sodium dodecyl sulfate polyacrylamide gel electrophoresis, and a pH and temperature optima of 4.0 and 50 °C. With nitroblue tetrazolium and riboflavin as substrates, the K _m values were 57.31 ± 0.012 and 1.51 ± 0.014 M, respectively, with corresponding V _max values of 333.7 ± 0.034 and 254.1 ± 0.022 μmol min⁻¹ mg protein⁻¹. This SOD likely belongs to the Fe- or Mn-SOD category due to the fact that it was insensitive to potassium cyanide or hydrogen peroxide inhibition, but was potentially weakly stimulated by hydrogen peroxide, and stimulated by Mn²⁺and Fe²⁺ ions. Moreover, this purified SOD also exhibited inhibitory effects on lipopolysaccharide-induced nitric oxide production in cultured mouse macrophage cell RAW 264.7 in a dose-dependent manner (IC₅₀ = 14.36 ± 0.15 μg protein/ml).     

Production of Lipids Containing High Levels of Docosahexaenoic Acid by a Newly Isolated Microalga, <Emphasis Type="Italic">Aurantiochytrium</Emphasis> sp. KRS101

In the present study, a novel oleaginous Thraustochytrid containing a high content of docosahexaenoic acid (DHA) was isolated from a mangrove ecosystem in Malaysia. The strain identified as an Aurantiochytrium sp. by 18S rRNA sequencing and named KRS101 used various carbon and nitrogen sources, indicating metabolic versatility. Optimal culture conditions, thus maximizing cell growth, and high levels of lipid and DHA production, were attained using glucose (60 g l⁻¹) as carbon source, corn steep solid (10 g l⁻¹) as nitrogen source, and sea salt (15 g l⁻¹). The highest biomass, lipid, and DHA production of KRS101 upon fed-batch fermentation were 50.2 g l⁻¹ (16.7 g l⁻¹ day⁻¹), 21.8 g l⁻¹ (44% DCW), and 8.8 g l⁻¹ (40% TFA), respectively. Similar values were obtained when a cheap substrate like molasses, rather than glucose, was used as the carbon source (DCW of 52.44 g l⁻¹, lipid and DHA levels of 20.2 and 8.83 g l⁻¹, respectively), indicating that production of microbial oils containing high levels of DHA can be produced economically when the novel strain is used.     

Production and Characterization of Cellulose by <Emphasis Type="Italic">Acetobacter</Emphasis> sp. V6 Using a Cost-Effective Molasses–Corn Steep Liquor Medium

In order to reduce of the manufacturing cost of bacterial cellulose (BC), BC production by Acetobacter sp. V6 was investigated in shaking culture using molasses and corn steep liquor (CSL) as the sole carbon and nitrogen sources, respectively. The highest BC production was obtained with Ca₃(PO₄)₂-treated molasses. Maximum BC yield (2.21 ± 0.04 g/l) was obtained at 5% (w/v) total sugar in molasses. In improved medium containing molasses and CSL, BC production was observed in the medium after 1 day of incubation and increased rapidly thereafter with maximum yield (3.12 ± 0.03 g/l) at 8 days. This value was approximately twofold higher than the yield in the complex medium. Physical properties of BC from the complex and molasses media were studied using Fourier-transform infrared (FT-IR) spectroscopy and X-ray diffractometer. By FT-IR, all the BC were found to be of cellulose type І, the same as typical native cellulose. The relative crystallinity of BC produced in the complex and molasses media were 83.02 and 67.27%, respectively. These results suggest that molasses and CSL can be useful low-cost substrates for BC production by Acetobacter sp. V6 without supplementation with expensive nitrogen complexes such as yeast extract and polypeptone, leading to the reduction in the production costs.     

In Vitro Adventitious Shoot Regeneration via Indirect Organogenesis from Petiole Explants of Cassia angustifolia Vahl.—a Potential Medicinal Plant

An effective protocol was developed for in vitro regeneration of the Cassia angustifolia via indirect organogenesis from petiole explants excised from 21-day-old axenic seedlings. Organogenic callus were induced on Murashige and Skoog (MS) medium supplemented with 5.0 μM 2,4-dichlorophenoxy acetic acid and 2.5 μM thidiazuron (TDZ). Adventitious shoot regeneration was achieved on MS medium supplemented with 5.0 μM TDZ as it induced 8.5 ± 0.98 shoots in 85% cultures. The number of shoots and shoot length was significantly enhanced when cultures were subcultured on auxin–cytokinin-containing medium. The highest number of shoots (12.5 ± 1.10) and shoot length (4.3 ± 0.20 cm) was recorded on MS medium supplemented with 5.0 μM TDZ and 1.5 μM indole-3-acetic acid. Regenerated shoots were rooted best on MS medium supplemented with 10.0 μM indole-3-butyric acid followed by their transfer to liquid MS filter paper bridge medium. The plants were successfully hardened off in sterile soilrite followed by their establishment in garden soil with 70% survival rate. The plants showed normal morphological characteristics similar to the field grown plants.