Submerged fermentation in wheat substrates for production of Monascus pigments

Cereal grains are normally used as solid substrates for the production of Monascus metabolites. However, solid fermentation in these substrates requires complex control systems, whereas in liquid culture the control of the fermentation is simpler and consequently significant reductions in fermentation times can be achieved. In the same way, the use of submerged culture can benefit the production of many secondary metabolites and decrease production costs by reducing the labour involved in solid-state methods. A flour composed of a mixed variety of Canadian hard wheat was used as sole nutrient source to produce the pigments of Monascus purpureus Went (IMI 210765). Supplementation with NH₄Cl promoted biomass and orange dye formation, whereas the use of zinc sulphate favoured red dyes production. In submerged fermentations significant differences in final pigment yields were observed in the use of wheat-based broth at different concentrations in the presence of bran particles and/or gluten protein. It has been found that the viscosity of the broth had a significant effect on the growth morphology and production of pigments. Gluten-free wheat flour at concentrations of 3–5% was found to be the most suitable for liquid Monascus culture. The subsequent use of passive immobilization of Monascus served to enhance red pigment yields and to facilitate the downstream processing of the dyes.     

Repeated-batch production of pigments by immobilised Monascus purpureus

Extracellular pigment production by immobilised Monascus purpureus C322 has been studied in repeated-batch processes using different immobilising carriers such as Ca-alginate, polyurethane sponge, active carbon and pearlite. With Ca-alginate, pigment production was maximum (30.5 UA₄₇₀ as process mean production, three batches) while the cell leakage was negligible (0.4 g l⁻¹ free biomass) and the bead mechanical stability good; with this carrier, an extended repeated-batch fermentation (nine batches, 55 days) was carried out: the process pigment productivity was 3.87 UA₄₇₀ day⁻¹.     

Solid-state fermentation of carob pods for ethanol production

The production of ethanol from carob pods by Saccharomyces cerevisiae in solid-state fermentation was investigated. The maximal ethanol concentration (160±3 g/kg dry pods), ethanol productivity (6.7 ± 0.2 g/kg per hour), ethanol yield (40 ± 1.8%), biomass concentration (7.5 ± 0.4 x 10⁸ cells/g carob pulp) and fermentation efficiency (80 ± 2%) were obtained at an inoculum amount of 3%, a particle size of 0.5 mm, a moisture level of 70%, a pH of 4.5 and a temperature of 30°C. Under the same fermentation conditions both sterilized and non-sterilized carob pods pulp gave the same maximum ethanol concentration.     

Solid-state fermentation of grape pomace for ethanol production

Summary Grape pomace was used as a substrate for the production of ethanol under solid-state fermentation conditions. The yield of ethanol amounted to greater than 80% of the theoretical, based on the fermentable sugar consumed.     

Leucine interference in the production of water-soluble red Monascus pigments

The formation of soluble Monascus red pigments is strongly positively and negatively regulated by different amino acids. Leucine, valine, lysine, and methionine had strong negative effects on pigment formation. Leucine supported poor pigment formation when used as sole nitrogen source in fermentations, yet it neither repressed pigment synthase(s) nor inhibited its action. The new pigments derived from the hydrophobic leucine were more hydrophilic than the conventional red pigments (lacking an amino acid side-chain) and were extracellularly produced. Therefore, the low level of red pigments produced when leucine was the nitrogen source was not due to feed-back regulation by cell-bound leucine pigments. The negative effect of leucine was caused by enhanced decay of pigment synthase(s). The enhanced decay was not due simply to de novo synthesis of a leucine-induced protease.Abbreviations mSG Monosodium glutamate - MOPS 3-(N-morpholine)propane sulfonic acid - DCW dry cell weight     

Optimal C:N ratio for the production of red pigments by Monascus ruber

The carbon-to-nitrogen (C:N) ratio in the biomass of microfungi tends to be quite different (e.g. 10–15) compared with the C:N ratio in the red pigments (e.g. >20) of the fungus Monascus ruber. Therefore, determining an optimal C:N ratio in the culture medium for maximizing the production of the pigments is important. A culture medium composition is established for maximizing the production of the red pigment by the fungus M. ruber ICMP 15220 in submerged culture. The highest volumetric productivity of the red pigment was 0.023 AU L^?1 h^?1 in a batch culture (30 °C, initial pH of 6.5) with a defined medium of the following composition (g L^?1): glucose (10), monosodium glutamate (MSG) (10), MgSO₄·7H₂O (0.5), KH₂PO₄ (5), K₂HPO₄ (5), ZnSO₄·7H₂O (0.01), FeSO₄·7H₂O (0.01), CaCl₂ (0.1), MnSO₄·H₂O (0.03). This medium formulation had a C:N mole ratio of 9:1. Under these conditions, the specific growth rate of the fungus was 0.043 h^?1 and the peak biomass concentration was 6.7 g L^?1 in a 7-day culture. The biomass specific productivity of the red pigment was 1.06 AU g^?1 h^?1. The best nitrogen source proved to be MSG although four other inorganic nitrogen sources were evaluated.     

Optimization of submerged fermentation medium for citrinin-free monascin production by Monascus

Microbial fermentation of citrinin-free Monascus pigments is in favor in the development of food industry. This study investigated the influences of carbon source, nitrogen source, and mineral salts on the cell growth, monascin (MS), and citrinin (CT) production in Monascus M9. A culture medium composition was established for maximizing the production of citrinin-free MS in submerged culture, as follows: 50?g/L Japonica rice powder, 20?g/L NH₄NO₃, 3?g/L NaNO₃, 1.5?g/L KH₂PO₄, 1?g/L MgSO₄?·?7H₂O, 0.2?g/L MnSO₄. Under these conditions, no CT was detectable by high performance liquid chromatography. The yield of MS reached 14.11?mg/g, improving approximately 30% compared with before optimization.     

里氏木霉306产t-PA固态发酵条件的研究

对基因工程菌株里氏木霉(Trichodermareesei)306生物合成组织型纤溶酶原激活剂(t-PA)的固态发酵培养基成分和发酵条件进行了研究;分析了发酵过程中t-PA的生成、总糖的消耗和菌体生长的规律。在优化的固体发酵条件下,t-PA的最大酶活是924.63IU/g干重培养基,较初始条件下提高了5倍。通过浅盘进行了放大实验,最大酶活为983.64IU/g干重培养基,t-PA合成速率为:13.66IU/g干重培养基.h,较三角瓶固态发酵最高产酶提高了6.38%,产酶速率提高了24.07%。     

Solid-state fermentation: a continuous process for fungal tannase production

Truly continuous solid-state fermentations with operating times of 2-3 weeks were conducted in a prototype bioreactor for the production of fungal (Penicillium glabrum) tannase from a tannin-containing model substrate. Substantial quantities of the enzyme were synthesized throughout the operating periods and (imperfect) steady-state conditions seemed to be achieved soon after start-up of the fermentations. This demonstrated for the first time the possibility of conducting solid-state fermentations in the continuous mode and with a constant noninoculated feed. The operating variables and fermentation conditions in the bioreactor were sufficiently well predicted for the basic reinoculation concept to succeed. However, an incomplete understanding of the microbial mechanisms, the experimental system, and their interaction indicated the need for more research in this novel area of solid-state fermentation.     

Protease inhibitors from jackfruit seed (Artocarpus integrifolia)

Protease inhibitory activity in jackfruit seed (Artocarpus integrifolia) could be separated into 5 fractions by chromatography on DEAE-cellulose at pH 7.6. A minor fraction (I) that did not bind to the matrix, had antitryptic, antichymotryptic and antielastase activity in the ratio 24:1.9:1.0. Fraction II bound least tightly to the ion exchanger eluting with 0.05 M NaCl and could be resolved into an elastase/chymotrypsin inhibitor and a chymotrypsin/trypsin inhibitor by chromatography on either immobilized trypsin or phenyl Sepharose CL-4B. Fractions III and IV eluted successively with 0.10 M NaCl and 0.15 M NaCl from DEAE-cellulose, inhibited elastase, chymotrypsin and trypsin in the ratio 1.0: 0.53:0.55 and 1.0:8.9:9.8 respectively. Fraction V, most strongly bound to the matrix eluting with 0.3 M NaCl and was a trypsin/chymotrypsin inhibitor accounting for 74% of total antitryptic activity. This inhibitor was purified further. The inhibitor with a molecular weight of 26 kd was found to be a glycoprotein. Galactose, glucose, mannose, fucose, xylose, glucosamine and uronic acid were identified as constitutent units of the inhibitor. Dansylation and electrophoresis in the presence of mercaptoethanol indicated that the inhibitor is made up of more than one polypeptide chain. The inhibitor combined with bovine trypsin and bovine α-chymotrypsin in a stoichiometric manner as indicated by gel chromatography. It had very poor action on subtilisin BPN′, porcine elastase, pronase,Streptomyces caespitosus protease andAspergillus oryzae protease. It powerfully inhibited the caseinolytic activities of rabbit and horse pancreatic preparations and was least effective on human and pig pancreatic extracts. Modification of amino groups, guanido groups and sulphydryl groups of the inhibitor resulted in loss of inhibitory activity. Reduction of disulphide bridges, reduction with sodium borohydride and periodate oxidation also decreased the inhibitory activity.     

Solid-state fermentation: a promising microbial technology for secondary metabolite production

Solid state (substrate) fermentation (SSF) has been used successfully for the production of enzymes and secondary metabolites. These products are associated with the stationary phase of microbial growth and are produced on an industrial scale for use in agriculture and the treatment of disease. Many of these secondary metabolites are still produced by submerged liquid fermentations (SmF) even though production by this method has been shown to be less efficient than SSF. As large-scale production increases further, so do the costs and energy demands. SSF has been shown to produce a more stable product, requiring less energy, in smaller fermenters, with easier downstream processing measures. In this article we review an important area of biotechnology, since the recent evidence indicates that bacteria and fungi, growing under SSF conditions, are more than capable of supplying the growing global demand for secondary metabolites.     

Azaphilone pigments from a yellow mutant of the fungus Monascus kaoliang

Azaphilone pigments, monascusones A (1) and B (2), together with two known azaphilones, monascin (3) and FK17-P2b2 (4), were isolated from the CH2Cl2 extract of a yellow mutant of the fungus M. kaoliang grown on rice. Structures of the isolated compounds were elucidated by analyses of spectroscopic data. Monascusone A (1), the major metabolite of M. kaoliang, showed no antimalarial (against Plasmodium falciparum), antitubercular (against Mycobacterium tuberculosis H37Ra), and antifungal (toward Candida albicans) activities. Compound 1 exhibited no cytotoxicity against BC (breast cancer) and KB (human epidermoid carcinoma of cavity) cell lines.     

Production of water-soluble yellow pigments via high glucose stress fermentation of Monascus ruber CGMCC 10910

Monascus pigments are secondary metabolites of Monascus species and are mainly composed of yellow pigments, orange pigments and red pigments. In this study, a larger proportion of Monascus yellow pigments could be obtained through the selection of the carbon source. Hydrophilic yellow pigments can be largely produced extracellularly by Monascus ruber CGMCC 10910 under conditions of high glucose fermentation with low oxidoreduction potential (ORP). However, keeping high glucose levels later in the culture causes translation or a reduction of yellow pigment. We presume that the mechanism behind this phenomenon may be attributed to the redox level of the culture broth and the high glucose stress reaction of M. ruber CGMCC 10910 during high glucose fermentation. These yellow pigments were produced via high glucose bio-fermentation without citrinin. Therefore, these pigments can act as natural pigments for applications as food additives.

     

Solid-state fermentation for production of griseofulvin on rice bran using Penicillium griseofulvum

Griseofulvin is a secondary metabolite produced from fungal species that have morphology suitable for solid-state fermentation (SSF). Reports on production of griseofulvin by SSF are scarce. The present work investigates SSF for griseofulvin production, optimization of its process parameters vis-à-vis the conventional submerged fermentation and its downstream processing from the same. Rice bran adjusted to an initial moisture content (IMC) of 50% (v/w) inoculated with 1 mL of a suspension of 10(6) spores/mL under agitation at 250 rpm containing the modified Czapek-Dox medium and additional 0.1% choline chloride as a precursor gave a yield of griseofulvin in 9 days that was comparable to submerged fermentation after 28 days. The yield of griseofulvin (microg/g dry biomass) was comparable in SSF and submerged fermentation. The biomass was estimated by estimation of chitin. Discussions on the effect of each parameter in SSF have also been included.     

青霉PT95菌固态发酵产生类胡萝卜素的研究

本文对青霉Ｐｅｎｉｃｉｌｌｉｕｍｓｐ．ＰＴ９５菌株在固态发酵条件下菌核内产生类胡萝卜素进行了初步研究。结果表明,在３种固态发酵培养基中,玉米粉培养基（ＳＭＡ）比麸皮２基和棉籽壳培养基更适合于ＰＴ９５菌株固态发酵产生类胡萝卜素。为了增加菌核干重和提高类胡萝卜素产率,ＳＭＡ中需要添加氮源、碳源和植物油。在所度的各种氮、碳源中,以硝酸钠和麦芽糖效果最佳。通过我试验确定了在培养基盐溶液中添加硝酸钠３ｇ／Ｌ     

Solid-state fermentation for xylanase production by Thermoascus aurantiacus using response surface methodology

We investigated xylanase production by Thermoascus aurantiacus using semisolid fermentation. Multivariant statistical approaches were employed to evaluate the effects of several variables (initial moisture in the medium, cultivation time, inoculum level, and bagasse mass) on xylanase production. The initial moisture content and bagasse mass were the most important factors affecting xylanase activity. The xylanase activity produced by the fungus under the optimized conditions (81% moisture content and 17 g bagasse) was found to be 2700 U per gram of initial dry matter, whereas its value predicted by a polynomial model was 2400 U per gram of initial dry matter. Received: 4 December 1998 / Received revision: 15 March 1999 / Accepted: 16 May 1999     

Effect of pH on citrinin and red pigments production by Monascus purpureus CCT3802

The production of red pigments and citrinin by Monascus purpureus CCT3802 was investigated in submerged batch cultures performed in two phases: in the first phase, cells were grown on glucose, at pH 4.5, 5.5 or 6.5; after glucose depletion, pH was adjusted, when necessary, to 4.5, 5.5, 6.5, 7.0, 8.0 or 8.5, for a production phase. The highest total red pigments absorbance of 11.3 U was 16 times greater than the lowest absorbance and was achieved with growth at pH 5.5, followed by production at pH 8.5, which causes an immediate reduction of the intra cellular red pigments from 75% to 17% of the total absorbance. The lowest citrinin concentration, 5.5 mg L⁻¹, was verified in the same culture while the highest concentration, 55 mg L⁻¹, was verified in cultures entirely carried out at pH 5.5. An alkaline medium, besides promoting intra cellular red pigments excretion, strongly represses citrinin synthesis.     

Solid-state fermentation of sugar-beet

Summary Semi solid culture is adaptated to ethanol production from sugar-beet. This technique offers some advantages (reduction of liquid volumes and suppression of the diffusion step). With a tubular reactor, the ethanol concentration obtained reaches 8,3%, and the yield is 0,407. The feasability of solid state fermentation on sugar-beet is demonstrated.