Philippe J. Blanc

Production of citrinin by various species ofMonascus

SummaryThe production of citrinin by variousMonascus species was determinated using various culture mediums and conditions. The maximal production was obtained in fermentor usingM. ruber with concentrations of 380 mg/l. Since citrinin is a toxic product, it is essential that the production of red pigments as food additives fromMonascus sp. avoid the occurrence of citrinin; so, we argue that some nitrogen sources are unfavorable to the production of citrinin.

Kinetic analysis of red pigment and citrinin production by Monascus ruber as a function of organic acid accumulation.

In submerged cultures performed in synthetic medium containing glucose and glutamate, the filamentous fungus Monascus ruber produced a red pigment and a mycotoxin, citrinin. In oxygen-limiting conditions, the production of these two metabolites was growth-associated, as was the production of primary metabolites. In oxygen-excess conditions, the profile of citrinin production was typical of a secondary metabolite, since it was produced mostly during the stationary phase. In contrast, the production of the pigment decreased rapidly throughout the culture, showing a profile characteristic of an inhibitory mechanism. The organic acids produced during the culture, L-malate and succinate, were shown to be slightly inhibitory against pigment production, while citrinin production was unaffected. However, this inhibition could not account for the observed profile of pigment production in batch cultures. Other dicarboxylic acids such as fumarate or tartrate showed a similar effect to that provoked by malate and succinate as regards pigment production. It was concluded that the decrease in red pigment production during the culture was due to the inhibitory effect of an unknown product whose accumulation was favored in aerobic conditions.

Influence of nitrogen and iron limitations on lipid production by Cryptococcus curvatus grown in batch and fed-batch culture

Abstract Cryptococcus curvatus , an oleaginous yeast, was grown under iron (Fe) and nitrogen (N)-limited conditions to produce lipid with a high stearic acid (C18:0) content in batch and fed-batch culture. In batch culture, lack of ferrous ions had no evident effect on growth, lipid production or lipid composition. The highest stearic acid content in the total lipid was 14% in N-limited medium in batch culture. The combined effect of Fe and N-limitation in fed-batch culture was to increase the stearic acid content to 19% of the total lipids and the consumed carbon to nitrogen (C:N) ratio to 20 g/g. The highest biomass concentration achieved was 70 g/litre and 53% (w/w) lipid accumulated when grown for about 172 h in fed-batch culture. A C:N ratio between 20 and 50 was optimal for intracellular lipid accumulation and a ratio > 50 g/g became crucial for both growth and lipid production in fed-batch culture. The maximal C:N ratio achieved at the end of the culture was 75 g/g. Lipid yield from glucose (g lipid/g glucose) was 0·14 and the fatty acids of lipids obtained were mainly oleic (C18:1), palmitic (C16:0) and stearic (C18:0).

Characterization of the tea fungus metabolites

SummaryThe tea fungus (commonly designed as “kombucha”) is a symbiotic culture of at least three microorganisms: the acetic acid bacteria Acetobacter xylinum and two yeasts Zygosaccharomyces rouxii and Candida sp. in sugared tea (Hesseltine, 1965; Anonymous, 1983). These microorganisms were cultured in their traditional medium and several metabolites were identified and quantified : ethanol, lactic, acetic, gluconic and glucuronic acids. The antibacterial product known as usnic acid was also searched.

Medium-Chain Fatty Acids Affect Citrinin Production in the Filamentous Fungus Monascus ruber

ABSTRACT During submerged culture in the presence of glucose and glutamate, the filamentous fungus Monascus ruber produces water-soluble red pigments together with citrinin, a mycotoxin with nephrotoxic and hepatoxic effects on animals. Analysis of the13C-pigment molecules from mycelia cultivated with [1-13C]-, [2-13C]-, or [1,2-13C]acetate by 13C nuclear magnetic resonance indicated that the biosynthesis of the red pigments used both the polyketide pathway, to generate the chromophore structure, and the fatty acid synthesis pathway, to produce a medium-chain fatty acid (octanoic acid) which was then bound to the chromophore by atrans-esterification reaction. Hence, to enhance pigment production, we tried to short-circuit the de novo synthesis of medium-chain fatty acids by adding them to the culture broth. Of fatty acids with carbon chains ranging from 6 to 18 carbon atoms, only octanoic acid showed a 30 to 50% stimulation of red pigment production, by a mechanism which, in contrast to expectation, did not involve its direct trans-esterification on the chromophore backbone. However, the medium- and long-chain fatty acids tested were readily assimilated by the fungus, and in the case of fatty acids ranging from 8 to 12 carbon atoms, 30 to 40% of their initial amount transiently accumulated in the growth medium in the form of the corresponding methylketone 1 carbon unit shorter. Very interestingly, these fatty acids or their corresponding methylketones caused a strong reduction in, or even a complete inhibition of, citrinin production byM. ruber when they were added to the medium. Several data indicated that this effect could be due to the degradation of the newly synthesized citrinin (or an intermediate in the citrinin pathway) by hydrogen peroxide resulting from peroxisome proliferation induced by medium-chain fatty acids or methylketones.

Production of 2-Phenylethyl Alcohol by Kluyveromyces marxianus

Among the numerous yeasts able to produce flavor compounds, several strains of Kluyveromyces species are known for the synthesis of large amounts of aromatic compounds. In particular, the accumulation of 2‐phenylethyl alcohol in liquid cultures of Kluyveromyces marxianus was observed after incubation on a semisynthetic medium at 30 °C, 125 rpm. Changes in the carbon sources led to a significant modification of the amount of this aromatic compound; moreover, the variations in the level of dl‐phenylalanine and the exclusive use of a precursor isomer (l) also affected the synthesis of 2−phenylethyl alcohol. Finally, this study revealed the growth inhibition of K. marxianus when the concentration of 2‐phenylethyl alcohol reached a critical value near 1.4 g/L; the use of an exponential model allowed the inhibition constant ki to be determined (ki = 0.35 g/L) .

Lipid production by an unsaturated fatty acid auxotroph of the oleaginous yeast Apiotrichum curvatum grown in single-stage continuous culture

An unsaturated fatty acid auxotroph of the oleaginous yeast Apiotrichum curvatum, named UfaM3, blocked in the conversion of stearic to oleic acid was cultivated in single-stage continuous culture. The influence of consumed carbon to nitrogen ratios (C/N ratios, g g−1) obtained at various dilution rates (D) on fatty acid (FA) accumulation and its profiles were studied. In continuous culture in N-limited medium a maximum FA accumulation of 45.6% (g g−1 of dry biomass) was obtained at an optimal D of 0.049 h−1, recording an efficiency of substrate conversion of 0.48 g g−1 and 0.22 g g−1 for biomass and lipids, respectively. The quality of lipid approached cocoa butter at an optimal C/N ratio of between 20 and 30. The C/N ratio in the incoming medium was 38.5 g g−1 with 30 g l−1 of glucose and both C and N sources were completely consumed at a critical D of ≤ 0.07 h−1. The stability of the mutant was demonstrated in the steady-state conditions of the chemostat with regard to the FA composition of its lipids.

Growth kinetics and astaxanthin production of Phaffia rhodozyma on glycerol as a carbon source during batch fermentation

Glycerol was studied as a substrate for astaxanthin by Phaffia rhodozyma PR 190. With co-utilisation of yeast extract and peptone, the maximum specific growth rate was 0.24 ± 0.02 h−1. Astaxanthin percentage in total pigment is constant (0.78 mg/g) and its yield from glycerol is always 0.97 mg/g. The yield of biomass from glycerol alone is 0.50 ± 0.02 g/g. The specific rate of astaxanthin production versus the cell growth rate reached a maximum for an optimal specific growth rate of 0.075 h−1. For this optimal value, the maximum specific astaxanthin production rate is 0.09 ± 0.01 mg/g.h. The best astaxanthin results were : 33.7 mg/l, 0.2 mg/l.h and 1.8 mg/g yeast after a fermentation term of 168 hours. Our results suggest a strategy of astaxanthin production in fed batch culture or chemostat at a growth rate of 0.075 h−1.

Survival of alginate-entrapped cells of Azospirillum lipoferum during dehydration and storage in relation to water properties

Survival of alginate-entrapped cells of Azospirillum lipoferum was studied during dehydration using a dry air stream and during prolonged storage at various constant water activity values (aw). During the drying operation, the viability loss remained almost constant from the initial water content to 0.35 g water/g dry weight (DW) corresponding to a 98.5% water removal, strongly increased until a water content of 0.25 g/g DW and then stopped until the end of the drying operational (final aw 0.18). A water content of 0.25 g/g DW (aw=0.55) corresponded to the critical point of the moisture sorption isotherm curve from which water became restricted to the dry material. A high drying rate (5 g/g DW per hour) was shown to be more detrimental for cell viability than a low drying rate (1.18 g/g DW per hour). When the product was stored in a closed chamber with a regulated aw (0.23), the number of living cells decreased during a short period (less than 15 days) corresponding to the product aw stabilization, and then remained constant for more than 150 days. In addition, cell survival during storage was not affected by aw values in the range 0–0.55. Above aw=0.55, the higher the aw and the storage duration, the lower the residual survival percentage. The influence of the drying and storage conditions on the cell death rate are discussed with regard to both the mechanisms generally involved in viability loss and the hydration properties of water.

Propionic acid and biomass production using continuous ultrafiltration fermentation of whey

SummaryThis paper presents a study of propionic acid and propionibacteria production from whey by usingPropionibacterium acidi-propionici in continuous fermentation with cell recycle. The highest propionic acid volumetric productivity achieved was 5 g.l−1.h−1 with no biomass bleeding. A maximal biomass concentration of 130 g.l−1 was achieved before initiating biomass bleeding to give a biomass volumetric productivity of 3.2 g.l−1.h−1 with a biomass of 75 g.l−1 and a propionic acid productivity of 3.6 g.l−1.h−1 (for about 100 hours i.e. more than 50 residence times).