Elsevier

Bioresource Technology

Volume 76, Issue 3, February 2001, Pages 283-286
Bioresource Technology

Short communication
γ-Linolenic acid production by solid-state fermentation of Mucorales strains on cereals

https://doi.org/10.1016/S0960-8524(00)00097-3Get rights and content

Abstract

Oleaginous fungi of the genus Mucorales were screened for γ-linolenic acid (GLA) production on solid substrates containing moistened cereals. Cunninghamella elegans CCF 1318 produced the highest yields of GLA when cultivated on barley. Substrate moisture and cultivation temperature proved critical for effective GLA production. Vegetable oil supplied to the cultures improved GLA production. Rotating bottles and plastic bags were used as cultivation vessels to reproduce the conditions found in rotating drums and tray bioreactors, respectively. After 11 days of cultivation at 21°C, C. elegans produced 14.2 mg of GLA per gram of dry substrate, composed of a mixture of barley, spent malt grains (SMG) and peanut oil. GLA represented 15.6% of the total fatty acids in the lipid extract.

Introduction

γ-Linolenic acid (GLA, cis,cis,cis-6,9,12-octadecatrienoic acid), is an intermediate in the transformation of linolenic acid into prostaglandins, which play an important physiological role in the organism. In certain pathological conditions, such as diabetes, cancer, virus infection, as well as ageing, the enzyme Δ-6 desaturase that catalyses the conversion of linoleic acid into GLA is less active, thus leading to deficient prostaglandins synthesis. Furthermore, GLA deficiency has been associated with some skin disorders (Horrobin, 1992). This dietary supply of GLA has been suggested as useful to prevent these situations, but GLA is very rare in common food, so that specific sources of this fatty acid need to be provided. The GLA used in pharmacological preparations is generally obtained from the seed oil of several plants such as evening primrose, borage and blackcurrants. The first industrial process for the production of GLA by cultivation of oleaginous fungi was developed in the UK using Mucor circinelloides (Ratledge, 1993). Despite the good yields and quality of the product, the process proved unprofitable, due to marketing difficulties. Currently, a biotechnological process is successfully applied on the industrial scale in Japan using GLA producers of the genus Mortierella and Mucor. A further improvement in productivity was recently obtained growing a new mutant of Mortierella ramanniana in a Maxblend fermentor (Certik and Shimizu, 1999). Other fungi of the order Mucorales have been reported as promising GLA producers and various substrates and fermentation techniques have been applied in the attempt to enhance GLA production and make the fermentation process competitive with the traditional extraction from plant sources. In particular, fermentation on various solid substrates such as cereals (Emelyanova, 1996) and apple pomace (Stredansky et al., 2000) may be of potential use on the industrial scale. Solid-state fermentation (SSF) offers several advantages over the more commonly applied liquid submerged fermentation (LSF). Solid fermentation techniques entail low capital cost and low energy expenditure. The solid substrates are normally obtained at a low price as raw products or residues of agro and food-industries, yet they provide a concentrated source of most nutrients required for microbial growth and metabolite production. Furthermore, the environmental impact of the SSF process is usually lower than for LSF, since low wastewater is produced in the former, while the exhausted biomass, rich in assimilable proteins, can often be used to feed farm animals. In this work, various cereals were supplied as nutrient substrates to a number of fungal strains of the Mucorales, that were screened for GLA production. The fermentation process was optimised using the producing strain Cunninghamella elegans CCF 1318.

Section snippets

Methods

Micro-organisms maintenance and inoculum preparation. The fungal strains screened for GLA production are listed in Table 1. Stock cultures were maintained on Czapek–Dox agar slants at 4°C. The spore suspension used as inoculum was prepared by washing the mycelium, grown on Sabouraud agar for 4 days at 28°C, with a sterile aqueous solution of Tween 80 (0.1% w/v), obtaining ca. 1×105 spores/ml.

Cultivation in Erlenmeyer flasks. For strain screening and substrate selection, 500 ml Erlenmeyer flasks

Results and discussion

Screening of strains and substrate selection. Fungal strains of the order Mucorales were screened for GLA production on solid substrate consisting of pearl barley impregnated with a nutrient solution. Cunninghamella elegans CCF 1318 yielded the highest amounts of oil, with a favourable GLA content (Table 1), and was used in cultivations with various cereal substrates, as well as with combinations of cereals and oil (Table 2). Pearl barley cultivations resulted in the highest yields of fungal

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