Improvement of Omega-3 Docosahexaenoic Acid Production by Marine Dinoflagellate Crypthecodinium cohnii Using Rapeseed Meal Hydrolysate and Waste Molasses as Feedstock

Rapeseed meal and waste molasses are two important agro-industrial by-products which are produced in large quantities. In this study, solid state fermentation and fungal autolysis were performed to produce rapeseed meal hydrolysate (RMH) using fungal strains of Aspergillus oryzae, Penicillium oxalicum and Neurospora crassa. The hydrolysate was used as fermentation feedstock for heterotrophic growth of microalga Crypthecodinium cohnii that produce docosahexaenoic acid (DHA). The addition of waste molasses as a supplementary carbon source greatly increased the biomass and DHA yield. In the batch fermentations using media composed of diluted RMH (7%) and 1-9% waste molasses, the highest biomass concentration and DHA yield reached 3.43 g/L and 8.72 mg/L, respectively. The algal biomass produced from RMH and molasses medium also had a high percentage of DHA (22-34%) in total fatty acids similar to that of commercial algal biomass. RMH was shown to be rich in nitrogen supply comparable to the commercial nitrogen feedstock like yeast extract. Using RMH as sole nitrogen source, waste molasses excelled other carbon sources and produced the highest concentration of biomass. This study suggests that DHA production of the marine dinoflagellate C. cohnii could be greatly improved by concomitantly using the cheap by-products rapeseed meal hydrolysate and molasses as alternative feedstock.     

High gravity fermentation of sugarcane molasses to produce ethanol: Effect of nutrients

Fermentation efficiency of more than 85% was obtained by high gravity fermentation of 33–34°Bx (spec. gravity ≈1.134) molasses medium with certain nutrients, instead of generally employed medium containing ≈16% (w/v) total sugar (spec. gravity ≈1.090) for ethanol fermentation in distilleries to get maximum 80–85% conversion. The fermenting yeast, Saccharomyces, has varied capabilities, depending on the species and nutrition for fermenting the high solids medium. The fermentation period was reduced to 48 h; and the byproducts obtained were less in concentration, upon supplementation with nutrients (or osmoprotectants) like soy flour/wheat bran to the medium in the existing batch fermentation technology. This has been found partly due to improved yeast cell viability during fermentation.     

A cost-effective cane molasses medium for enhanced cell-bound phytase production by Pichia anomala

AIM: Formulation of an inexpensive cane molasses medium for improved cell-bound phytase production by Pichia anomala. METHODS AND RESULTS: Cell-bound phytase production by Pichia anomala was compared in synthetic glucose-beef extract and cane molasses media. The yeast was cultivated in 250 ml flasks containing 50 ml of the medium, inoculated with a 12 h-old inoculum (3 x 10(6) CFU ml(-1)) and incubated at 25 degrees C for 24 h at 250 rev min(-1). Different cultural parameters were optimized in cane molasses medium in batch fermentation. The cell-bound phytase content increased significantly in cane molasses medium (176 U g(-1) dry biomass) when compared with the synthetic medium (100 U g(-1) dry biomass). In fed-batch fermentation, a marked increase in biomass (20 g l(-1)) and the phytase yield (3000 U l(-1)) were recorded in cane molasses medium. The cost of production in cane molasses medium was pound 0.006 per 1000 U, which is much lower when compared with that in synthetic medium (pound 0.25 per 1000 U). CONCLUSIONS: An overall 86.6% enhancement in phytase yield was attained in optimized cane molasses medium using fed-batch fermentation when compared with that in synthetic medium. Furthermore, the production in cane molasses medium is cost-effective. SIGNIFICANCE AND IMPACT OF THE STUDY: Phytase yield was improved in cane molasses when compared with the synthetic medium, and the cost of production was also significantly reduced. This enzyme can find application in the animal feed industry for improving the nutritional status of feed and combating environmental pollution.     

Citric acid production by solid state fermentation using sugarcane bagasse

A solid state fermentation (SSF) method was used to produce citric acid by Aspergillus niger DS 1 using sugarcane bagasse as a carrier and sucrose or molasses based medium as a moistening agent. Initially bagasse and wheat bran were compared as carrier. Bagasse was the most suitable carrier, as it did not show agglomeration after moistening with medium, resulting in better heat and mass transfer during fermentation and higher product yield. Different parameters such as moisture content, particle size, sugar level and methanol concentration of the medium were optimised and 75% moisture level, 31.8 g sugar/100 g dry solid, 4% (v/w) methanol and particles of the size between 1.2 and 1.6 mm were found to be optimal. Sucrose and clarified and non-clarified molasses medium were also tested as moistening agents for SSF and under optimised conditions, 20.2, 19.8 and 17.9 g citric acid /100 g of dry solid with yield of 69.6, 64.5 and 62.4% (based on sugar consumed) was obtained in sucrose, clarified and non-clarified molasses medium respectively, after 9 days of fermentation.     

Influence of inocula and grains on sclerotia biomass and carotenoid yield of<Emphasis Type="Italic"> Penicillium</Emphasis> sp. PT95 during solid-state fermentation

Various inocula and grains were evaluated for carotenoid production by solid-state fermentation using Penicillium sp. PT95. Millet medium was more effective in both sclerotia growth and carotenoid production than other grain media. An inoculum in the form of sclerotia yielded higher sclerotia biomass compared to either a spore inoculum or a mycelial pellet inoculum. Adding wheat bran to grain medium favored the formation of sclerotia. However, neither the inoculum type nor addition of wheat bran resulted in a significant change in the carotenoid content of sclerotia. Among grain media supplemented with wheat bran (wheat bran:grain =1:4 w/w, dry basis), a medium consisting of rice and wheat bran gave the highest sclerotia biomass (15.10 g/100 g grain), a medium consisting of buckwheat and wheat bran gave the highest content of carotenoid in sclerotia (0.826 mg/g dry sclerotia), and a medium consisting of millet and wheat bran gave the highest carotenoid yield (11.457 mg/100 g grain).     

The fermentative production of oxytetracycline on industrial by-products byStreptomyces rimosus 12907

Three strains ofStreptomyces rimosus were grown on four different media. The one suitable for the production of oxytetracycline byStreptomyces rimosus 12907 was modified by black strap molasses, fodder yeast (40% total protein) and rice bran. The volume of the fermentation medium was scaled up in a 1200-litre fermentor aerated with sterile air obtained from a system used in the purification of air. 850 g crude oxytetracyeline was obtained when the fermented medium (700 litres) was extracted with 1-butanol. This work was carried out in the Laboratories of the Chemical Factories of the Egyptian Sugar anio Distillation Company with the cooperation of the National Research Centre, Dokki, Cairo, Arab Republd of Egypt (A.R.E.).     

Effect of carbon and nitrogen sources on growth and biological efficacy of Pseudomonas fluorescens and Bacillus subtilis against Rhizoctonia solani, the causal agent of bean damping-off

One of the most important environmental factors that regulate the growth and antagonistic efficacy of biocontrol agents is the medium. The aim of this paper was to find the nitrogen and carbon sources that provide maximum biomass production of strains P-5 and P-6 (Pseudomonas fluorescens), B-3 and B-16 (Bacillus subtilis) and minimum cost of media, whilst maintaining biocontrol efficacy. All of the strains were grown in seven liquid media (pH=6.9) including: sucrose + yeast extract, molasses of sugar beet + yeast extract in 2:1 and 1:1 w/w ratios, molasses of sugar beet + urea, nutrient broth, molasses and malt extract, at an initial inoculation of 1 x 10(5) CFU ml(-1). Cells from over night cultures used to inoculate soil at 1 x 10(9) CFU cm(-3) soil. At the same time, fungal inoculum (infected millet seed with Rhizoctonia solani) was added to soil at the rate of 2 g kg(-1) soil. Results indicated that growth of P-6, B-3 and B-16 in molasses + yeast extract (1:1 w/w) medium was significantly higher than in the other media. Molasses + yeast extract (1:1 and 2:1 w/w) media supported rapid growth and high cell yields in P-5. In greenhouse condition, results indicated that the influence of the media on the biocontrol efficacy of P-5, P-6, B-3 and B-16 was the same and Pseudomonas fluorescens P-5 in molasses and malt extract media reduced the severity of disease up to 72.8 percent. On the other hand, there were observed significant differences on bean growth after one month in greenhouse. P-5 in molasses + yeast extract (1:1 w/w) medium had the most effects on bean growth promotion. In this study molasses media showed good yield efficacy in all of the strains. The high sucrose concentration in molasses justifies the high biomass in all of the strains. Also, the low cost of molasses allows its concentration to be increased in media. On the other hand, yeast extract was the best organic nitrogen source for antagonist bacteria but it is expensive for an industrial process. So it should be replaced by another industrial product instead of yeast extract, which confirm by an economic and technological study. The results obtained in this study could be used to provide a reliable basis to increase the population of biocontrol agents in fermentation process.     

Solid-state fermentation for gluconic acid production from sugarcane molasses by Aspergillus niger ARNU-4 employing tea waste as the novel solid support

Solid-state fermentation (SSF) was evaluated to produce gluconic acid by metal resistant Aspergillus niger (ARNU-4) strain using tea waste as solid support and with molasses based fermentation medium. Various crucial parameters such as moisture content, temperature, aeration and inoculum size were derived; 70% moisture level, 30 degrees C temperature, 3% inoculum size and an aeration volume of 2.5l min(-1) was suited for maximal (76.3 gl(-1)) gluconic acid production. Non-clarified molasses based fermentation media was utilized by strain ARNU-4 and maximum gluconic acid production was observed following 8-12 days of fermentation cycle. Different concentrations of additives viz. oil cake, soya oil, jaggary, yeast extract, cheese whey and mustard oil were supplemented for further enhancement of the production ability of microorganism. Addition of yeast extract (0.5%) was observed inducive for enhanced (82.2 gl(-1)) gluconic acid production.     

Ergosterol production from molasses by genetically modified <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis>

Ergosterol is an economically important metabolite produced by fungi. Recombinant Saccharomyces cerevisiae YEH56(pHXA42) with increased capacity of ergosterol formation was constructed by combined overexpression of sterol C-24(28) reductase and sterol acyltransferase in the yeast strain YEH56. The production of ergosterol by this recombinant strain using cane molasses (CM) as an inexpensive carbon source was investigated. An ergosterol content of 52.6 mg/g was obtained with 6.1 g/l of biomass from CM medium containing 60 g/l of total sugar in 30 h in shake flask. The ergosterol yield was enhanced through the increasing cell biomass by supplementation of urea to a concentration of 6 g/l in molasses medium. Fermentation was performed in 5-l bioreactor using the optimized molasses medium. In batch fermentation, the effect of agitation velocity on ergosterol production was examined. The highest ergosterol yield was obtained at 400 rpm that increased 60.4 mg/l in comparison with the shake flask culture. In fed-batch fermentation, yeast cells were cultivated, firstly, in the starting medium containing molasses with 20 g/l of total sugar, 1.68 g/l of phosphate acid, and 6 g/l of urea (pH 5.4) for 5 h, then molasses containing 350 g/l of total sugar was fed exponentially into the bioreactor to keep the ethanol level in the broth below 0.5%. After 40 h of cultivation, the ergosterol yield reached 1,707 mg/l, which was 3.1-fold of that in the batch fermentation.     

Attempts at improving citric acid fermentation by Aspergillus niger in beet-molasses medium

Natural oils with high unsaturated fatty acids content when added at concentrations of 2% and 4% (v/v) to beet molasses (BM) medium caused a considerable increase in citric acid yield from Aspergillus niger. The fermentation capacities were also examined for production of citric acid using BM-oil media under different fermentation conditions. Maximum citric acid yield was achieved in surface culture in the presence of 4% olive oil after 12 days incubation.     

Cauliflower waste incorporation into cane molasses improves ethanol production using <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> MTCC 178

Diluted cane molasses having total sugar and reducing sugar content of 9.60 and 3.80% (w/v) respectively was subjected to ethanol production by Saccharomyces cerevisiae MTCC 178. Incorporation of dried Cauliflower Waste (CW) in molasses at the level of 15 % increased ethanol production by nearly 36 % compared to molasses alone. Addition of 0.2 % yeast extract improved ethanol production by nearly 49 % as compared to molasses alone. When the medium containing diluted molasses and 0.2 % yeast extract was supplemented with 15 % CW, 29 % more ethanol was produced compared to molasses with 0.2 % yeast extract. Cell biomass, ethanol production, final ethanol concentration and fermentation efficiency of 2.65 mg mL⁻¹, 41.2 gL⁻¹, 0.358 gg⁻¹ and 70.11 % respectively were found to be best at 15% CW supplementation level besides reduction in fermentation time but further increase in CW level resulted in decline on account of all the above parameters. This is probably the first report to our knowledge, in which CW was used in enhancing ethanol production significantly using a small quantity of yeast extract.     

绿色木霉在啤酒糟基质上产纤维素酶条件的优化

对绿色木霉接种到啤酒糟固态发酵产纤维素酶的培养基和培养条件进行优化,考察发酵物料起始含水量、发酵时间、起始pH值等发酵条件,以及啤酒糟培养基中添加麸皮、氮源种类对产酶的影响。结果表明,以啤酒糟为发酵基质接种绿色木霉生产纤维素酶是可行的。经单因素和正交试验获得最适固态发酵的培养条件为:起始pH 5~6,培养温度28~30℃,发酵4 d;最佳发酵培养基组合为:麸皮比例30%,培养基起始含水量50%,(NH₄)₂SO₄添加量为2.0%~2.5%。     

Protein enrichment of potato processing waste through yeast fermentation

Potato starch obtained from waste waters of chips manufacturing was used as a fermentation substrate for yeast protein enrichment. Among 18 yeast strains, 6 strains were screened according to their biomass yield and protein content after fermentation for 16 h at 30 degrees C in an aerated glucose-based liquid media (4.5 Ls). Using concentrated media (25% solids) made from potato starch pre-hydrolyzed with malt flour and batch-fermented for 20 h at 26 degrees C under aerobic conditions, Candida utilis ATCC 9256 was the most efficient protein-forming strain. Scaled-up at the 100 Ls level, the aerobic batch process was improved under fed-batch conditions with molasses supplementation. After drying, fermented starch contained 11-12% protein, including 7-8% yeast protein.     

Batch fermentation kinetics of sugar beet molasses by Zymomonas mobilis

A new osmotolerant mutant strain of Zymomonas mobilis was successfully used for ethanol production from beet molasses. Addition of magnesium sulfate to hydrolyzed molasses allowed repeated growth without the need of yeast extract addition. The kinetics and yields parameters of fermentation on media with different molasses concentrations were calculated. The anabolic parameters (specific growth rate, mu, and biomass yield, Y(X/S)) were inhibited at elevated molasses concentrations while the catabolic parameters (specific ethanol productivity, q(p), and ethanol yield, Y(p/s)) were not significantly affected. In addition to ethanol and substrate inhibition, osmotic pressure effects can explain the observed results.     

Optimization of medium composition for alkali-stable xylanase production by Aspergillus fischeri Fxn 1 in solid-state fermentation using central composite rotary design

Response surface methodology and central composite rotary design (CCRD) was employed to optimize a fermentation medium for the production of alkali-stable cellulase-free xylanase by Aspergillus fischeri in solid-state fermentation at pH 9.0 with wheat bran as substrate. The four variables involved in this study were sodium nitrite, potassium dihydrogen phosphate, magnesium sulphate and yeast extract. The statistical analysis of the results showed that, in the range studied, only sodium nitrite had a significant effect on xylanase production. The optimized medium containing (in g/l) NaNO(2)-7.0, K2HPO(4)-1.0, MgSO(4)-0.5 and yeast extract-5.0 resulted in 1.9-fold increased level of alkali-stable xylanase (1024 U/g wheat bran) production compared to initial level (540 U/g) after 72 h of fermentation, whereas its value predicted by the quadratic model was 931 U/g. The level of protease activity was considerably decreased in optimized medium, thus helping to preserve the xylanase activity and demonstrating another advantage of applying statistical experimental design.     

Conversion of oil waste to valuable fatty acids using Oleaginous yeast

Oil waste poses a highly dangerous threat to the environment, mainly because it is considered a high energy demanding degradation process. Oleaginous yeast utilizing oil waste to produce microbial fatty acids is considered an innovative method for oil waste elimination. In the present study, fifteen yeast isolates were screened for their lipid content, three of which were chosen for their high lipid content as compared to the standard strain Phaffia rhodozyma NRRL-Y-10921. The three selected isolates were further screened for their fatty acid profile. Yeast isolate (NC-I), identified as Yarrowia lipolytica, was chosen because it exceeded the lipid production of the standard strain by 21%, it also produced the highest C 14:0 (myristic acid), C 18:1 (oleic acid) and C18:2 (linoleic acid), compared to the other two isolates. Growth on different oil wastes resulted in an increase in total lipid content which reached its maximum when oil waste of frying vegetables was added to the media (57.89%). A variation in the fatty acid profile was detected when different types of oil waste were used before and after fermentation. The addition of different glucose concentrations to the vegetable oil waste media resulted in the appearance of C 22:0 (behenic acid) which was not present when the basal medium was used. Scanning Electron Microscopy indicated morphologic changes when the yeast was grown in high glucose concentration as compared to those grown in oil waste media. The activity of malate dehydrogenase (MDH) enzyme exhibited a correlative relationship with the lipid content under various glucose concentrations. The obtained results indicate that vegetable oil waste is suitable for microbial fatty acid production and that the fatty acid profile could be maneuvered through the manipulation of the fermentation media.     

Production of tyrosinase by Auricularia auricula using low cost fermentation medium

Low cost fermentation media using agricultural by-products (wheat bran extract, rice bran extract and soybean meal extract) as a major nutrient source, were evaluated for the production of tyrosinase from the fungus Auricularia auricula in submerged culture. In single-factor experiments, three components (wheat bran extract, casein and CuSO₄) were chosen to further optimize medium composition using response surface methodology (RSM). The central composite experimental results showed the following optimum medium composition: wheat bran extract 36.0 %, casein 1.1 g/l and CuSO₄ 0.13 g/l. Under these conditions, the highest tyrosinase activity was 17.22 U/ml, which was 2.1 fold higher than that obtained using the non-optimized medium. The present study is the first to report the statistical optimization of medium composition for production of tyrosinase by A. auricula using cheaper wheat bran extract as a major nutrient source. These results might provide a reference for the development of a cost-effective medium for commercial production of tyrosinase.     

Simultaneous saccharification and L-(+)-lactic acid fermentation of protease-treated wheat bran using mixed culture of lactobacilli

Protease-treated wheat bran (20% w/v) of particle size less than 300 μm containing 65% (w/w) starch was used for the simultaneous saccharification and l-(+)-lactic acid fermentation by the mixed cultures of Lactobacillus casei and Lactobacillus delbrueckii. Maximum lactate yield after various process optimizations was 123 gl⁻¹ with a productivity of 2.3 gl⁻¹ h⁻¹ corresponding to a conversion of 0.95 g lactic acid per gram starch after 54 h at 37°C. By using protease-treated wheat bran around tenfold decrease in supplementation of the costly medium component, like yeast extract, was achieved together with a considerable increase in the production level.     

High production system of the antibacterial peptide PLG-1

Propionibacterium thoenii P-127 produces and releases to the growth medium antibacterial agents that can be used as natural preservatives. The concentrations of these antibacterial agents in the growth medium are very low, and their activity can be detected only in concentrated medium, even in a bioreactor. A simple and efficient system to produce propionicin PLG-1 without the use of a bioreactor was investigated. Fermentation in screw-cap bottles without shaking produced antibacterial activity similar to that of fermentation in plates, but in a shorter time. Sodium lactate medium (NaLa) was found to be the most supportive for PLG-1 production compared to lactic acid bacteria media such as M-17 or beet molasses/corn. The initial concentration of the carbon source, sodium lactate, agar concentration, and the initial pH of the medium affected the synthesis of PLG-1. Additions of NaCl up to 1% showed no effect on the antibacterial agent production. The optimal conditions for production of the antibacterial agent were fermentation for 9 days in screw-cap bottles in modified NaLa medium (M-NaLa) containing 1% yeast extract, 1% tryptic soy broth, 0.9% lactic acid, and 0.6% agar, adjusted to pH of 9.     

Production of mosquitocidal <Emphasis Type="Italic">Bacillus sphaericus</Emphasis> by solid state fermentation using agricultural wastes

In this study, Bacillus sphaericus NRC 69 was grown in culture media, in which 12 agricultural wastes were tested as the main carbon, nitrogen and energy sources under solid state fermentation. Of the 12 tested agricultural by-products, wheat bran was the most efficient substrate for the production of B. sphaericus mosquitocidal toxins against larvae of Culex pipiens (LC₅₀ 1.2 ppm). Mixtures of tested agricultural wastes separately with wheat bran enhanced the produced toxicity several folds and decreased LC₅₀ between 3.7- and 50-fold in comparison with that of agricultural wastes without mixing. The toxicity of B. sphaericus grown in wheat bran/rice hull at 8/2 (g/g) and wheat bran/barley straw at 1/4 (g/g) showed the same toxicity as that in wheat bran medium (LC₅₀ decreased 17- and 16-fold, in comparison with that in rice hull or barely straw media, respectively). In wheat bran medium, the maximum toxicity of the tested organism obtained at 50% moisture content, inoculum size 84 × 10⁶ CFU/g wheat bran and incubation for 6 days at 30°C. Addition of cheese whey permeate at 10% to wheat bran medium enhanced the toxicity of B. sphaericus NRC 69 about 46%.