Production Of Γ-decalactone Research Articles

Effect of physical factors on the production of γ-decalactone by immobilized cells of Sporidiobolus salmonicolor

The effects of some physical factors, including preculture time, media initial pH and bead concentrations, on the production of γ-decalactone by immobilized Sporidiobolus salmonicolor CCRC 21975 within calcium alginate beads were investigated. Long-term reuse and stability of the immobilized preparation were also examined. Regardless of the preculture time, no significant difference in the production of γ-decalactone by immobilized S. salmonicolor cells was observed as long as the initial populations of the entrapped cells were the same. Calcium alginate-immobilized S. salmonicolor cells showed improved tolerance to perturbations in external pH and produced more γ-decalactone than did free cells. The patterns of changes in S. salmonicolor cells inside or outside the gel beads as well as production of γ-decalactone by immobilized S. salmonicolor did not vary significantly with bead concentration. Furthermore, after reuse of the calcium alginate-immobilized S. salmonicolor cells for 13 consecutive 4-day batch fermentations, no bead leakage was detected. The production of γ-decalactone in the 13th cycle was about 58.4% of that in the first cycle.

In situ detoxification of the fermentation medium during gamma-decalactone production with the yeast sporidiobolus salmonicolor

gamma-Decalactone (gamma-C10) is known to be highly toxic for the microorganims used for its production. In this work, three techniques were studied in order to overcome this toxicity during a bioconversion process using ricinoleic acid as precursor of the lactone: in situ trapping in oily phases, in porous hydrophobic sorbents and in beta-cyclodextrins. Oily phases added to the media (olive, Miglyol, tributyrin, and paraffin) had a protective effect on Sp. salmonicolor, and they improved the lactone production. beta-cyclodextrins, which have a hydrophobic cavity that can trap molecules such as gamma-C10, have been used to protect the yeasts. The results showed insufficient preservation of cell viability. Some sorbents (activated carbon and polystyrene-based sorbents) were successfully tested during bioconversion. In all cases viability exceeded the reference one. Nevertheless the aroma production was 30% lower than the reference. All of these solutions led to some enhancement of the cell viability during bioconversion of methyl ricinoleate to gamma-C10. For improvement of the lactone production, the oil trapping method seemed to be the best with the experimental conditions tested.

Peroxisomal beta-oxidation activities and gamma-decalactone production by the yeast Yarrowia lipolytica.

gamma-Decalactone is a peachy aroma compound resulting from the peroxisomal beta-oxidation of ricinoleic acid by yeasts. The expression levels of acyl-CoA oxidase (gene deletion) and 3-ketoacyl-CoA thiolase activities (gene amplification on replicative plasmids) were modified in the yeast Yarrowia lipolytica. The effects of these modifications on beta-oxidation were measured. Overexpression of thiolase activity did not have any effect on the overall beta-oxidation activity. The disruption of one of the acyl-CoA oxidase genes resulted in an enhanced activity. The enhancement led to an increase of overall beta-oxidation activity but reduced the gamma-decalactone production rates. This seemed to indicate a non-rate-limiting role for beta-oxidation in the biotransformation of ricinoleic acid to gamma-decalactone by the yeast Yarrowia lipolytica. All strains produced and then consumed gamma-decalactone. We checked the ability of the different strains to consume gamma-decalactone in a medium containing the lactone as sole carbon source. The consumption of the strain overexpressing acyl-CoA oxidase activity was higher than that of the wild-type strain. We concluded that peroxisomal beta-oxidation is certainly involved in gamma-decalactone catabolism by the yeast Y. lipolytica. The observed production rates probably depend on an equilibrium between production and consumption of the lactone.

Production of γ-decalactone from ricinoleic acid by immobilized cells of Sporidiobolus salmonicolor

Sporidiobolus salmonicolor CCRC 21975 was immobilized in κ-carrageenan, chitosan, agarose or calcium alginate. Due to the detrimental effects of high temperature attained during the gelling processes of κ-carrageenan and agarose as well as the toxicity of chitosan to the test organism, immobilization of S. salonicolor with these matrices for the production of γ-decalactone was inadequate. Neither viable cells nor production of γ-decalactone could be detected in media after 4 days cultivation of S. salmonicolor immobilized with κ-carrageenan or chitosan. Fewer viable cells and little γ-decalactone production was found in media with agarose-immobilized cells. In contrast, no significant reduction in the viable population was noted during immobilization procedures using alginate. Alginate-immobilized S. salmonicolor cells showed less susceptibility to ricinoleic acid toxicity and produced more γ-decalactone than did free cells. Time courses of γ-decalactone production by S. salmonicolor also revealed that immobilized cells produced a maximum γ-decalactone yield of ca. 131·8 mg l −1 after 5 days fermentation, compared with a maximum of ca. 107·5 mg l −1 for free cells.

Trapping of -Decalactone by Adsorption on Hydrophobic Sorbents : Application to the bioconversion of methyl ricinoleate by the yeast Sporidiobolus salmonicolor

Trapping by adsorption on hydrophobic porous polymers was used to selectively remove γ-decalactone from the complex bioconversion medium. Several sorbents were tested : activated carbon, and three porous polystyrene-type polymers (Porapak Q, Chromosorb 105, and Resin SM4). The sorption isotherms were determined in water and in a model bioconversion medium without microorganisms. In the last part of the study, the application to the production of γ-decalactone by Sporidiobolus salmonicolor was carried out. Hydrophobic porous adsorbents are compatible with the bioconversion and protect the yeast from the toxic effect of γ-decalactone.

Production of γ-decalactone and 4-hydroxy-decanoic acid in the genus Sporidiobolus

Within the genus Sporidiobolus, S. odorus has been widely reported as having the ability to convert castor oil or its derivatives to γ-decalactone, but other species have also shown potential for lactone production. In this work, the bioconversion of ricinoleic acid methyl ester to γ-decalactone was studied with four species of Sporidiobolus: S. salmonicolor, S. ruinenii, S. johnsonii, and S. pararoseus. With 4.1 g/ l of ricinoleic acid methyl ester, only S. salmonicolor and S. ruinenii were able to produce γ-decalactone (12 and 40 mg/ l respectively). S. johnsonii and S. pararoseus did not produce any lactone in spite of consuming the precursor. The four strains showed different sensitivities to lactone. Between S. salmonicolor and S. ruinenii, the latter was the best biocatalyst. During four successive batch cultivations in a 7- l bioreactor, 5.5 g/ l of γ-decalactone was produced with S. ruinenii in each 10-d run. This was essentially due to its ability to produce the open form of γ-decalactone ( i.e., 4-hydroxy-decanoic acid), which is far less toxic than the lactone.

Fatty acid accumulation in the yeast Sporidiobolus salmonicolor during batch production of γ-decalactone

This paper provides new information about the metabolism of various fatty acids and γ-decalactone production by yeast. An analysis of the fatty acid composition of the yeast Sporidiobolus salmonicolor during batch production of lactone with ricinoleic acid methyl ester as a precursor showed an accumulation of the γ-decalactone precursor inside the cells. Electron microscopy of the yeasts showed the presence of large internal inclusions leading to membrane and organelle lysis and, consequently, death of the yeast. S. salmonicolor cultivated with methyl oleate did not produce γ-decalactone and is viable during the whole culture. Analysis of the long chain fatty acid fraction showed incorporation of methyl oleate.

Effects of various fatty acid components of castor oil on the growth and production of γ-decalactone by Sporobolomyces odorus

The growth behavior of Sporobolomyces odorus in medium containing 0.06% palmitic, steraric or oleic acid was observed to be similar to that in medium lacking these acids. However, growth of S. odorus was partially inhibited in the presence of linoleic acid or ricinoleic acid. Addition of palmitic, stearic, oleic, or linoleic acid to the medium reduced the yields of γ-decalactone as determined after 168 h of cultivation. On the other hand, a significant increase in the yield of approximately 71.8 mg/ l, i.e. about 25 times that (3.5 mg/ l) in the control medium, was noted in the medium containing ricinoleic acid. The concentration of ricinoleic acid as well as the time of addition were also observed to affect the amount of γ-decalactone produced by S. odorus. When 0.06% ricinoleic acid was added to the medium at the beginning of cultivation, a maximum yield of approximately 135.4 mg/ l γ-decalactone was obtained after 216 h of cultivation.

Growth of and production of γ-decalactone by Sporobolomyces odorus in jar fermentors as affected by pH, aeration and fed-batch technique

In jar fermentors, Sporobolomyces odorus grew rapidly and entered the stationary growth phase at approximately 72 h of cultivation. During the 192 h fermentation period, pH and DO concentration in the medium first decreased and then increased. A maximum yield of γ-decalactone of 54.6 mg/ l was noted after 120 h of cultivation. Studies on the effects of pH control and aeration rate revealed that maintaining the pH of the medium at 4.0, 5.0, 6.0 or 7.0 neither affected the growth of S. odorus nor improved the yield of γ-decalactone, and that although a larger maximum final population was found in the culture with an aeration rate of 2 vvm, a greater yield of γ-decalactone was noted when aeration rate was maintained at 1.0 or 1.5 vvm. A significant increase in the maximum yield of γ-decalactone was noted in fed-batch culture of S. odorus. The maximum yield of γ-decalactone increased with the number of castor oil hydrolysate feedings. Fed-batch cultivation of S. odorus with additions of castor oil hydrolysate on the 3rd, 4th and 5th d of cultivation resulted in a maximum yield of 208 mg/ l γ-decalactone which was noted after 7 d of cultivation.

Growth and production of γ-decalactone and Cis-6-dodecen-4-olide by Sporobolomyces odorus in the presence of fatty acids and oils

Sporobolomyces odorus produces γ-decalactone and cis-6-dodecen-4-olide, substances which exhibit odors characteristic of peaches and lamb meat (mutton), respectively. The growth and production of γ-decalactone by S. odorus in broth to which fatty acids or oils had been added were investigated. The addition of decanoic acid or dodecanoic acid to the culture reduced the growth and production of γ-decalactone by S. odorus. Without affecting the growth, the presence of Miglyol oil reduced the yield, whereas the addition of castor oil enhanced the production of γ-decalactone by S. odorus. The greatest enhancement, with a yield of 8.62 mg/ l of γ-decalactone, about six times that in the control broth, was noted when 3% castor oil was added after 24 h of cultivation. On the other hand, the fatty acids and oils tested either reduced or had no effect on the production of cis-6-dodecen-4-olide by S. odorus.

Development of macrocapsules containing bioflavors generated in situ by immobilized cells

An alternative approach to microbial production of bioflavors, eliminating the need for lengthy product purification, is presented. It is based on co-immobilization of precursors for bioflavor generation by microbial cells, traditionally employed for food and beverage processing, within beads made of food-grade gel matrix. Following incubation under controlled conditions the bioflavor — or bioflavor mixture — is generated and accumulated within the beads. The flavor-retaining bead may then be employed as a food additive. A feasibility study demonstrated this approach with ethanol production by baker's yeast co-immobilized with glucose medium. Means and conditions for bead preparation and control of ethanol levels and production rate are presented. Complex bioflavor generation was also demonstrated by baker's yeast co-immobilized with apple juice, generating cider flavors. Beads providing beer taste were also readily made via co-immobilization of commercial brewing yeast with malt. Furthermore, the potential inherent in bioflavor generation by co-immobilization of filamentous fungi with an emulsion of oily precursor was demonstrated by γ-decalactone production from castor oil.

Presumptive involvement of methyl ricinoleate β-oxidation in the production of γ-decalactone by the yeast Pichia guilliermondii

Presumptive involvement of methyl ricinoleate β-oxidation in the production of γ-decalactone by the yeast Pichia guilliermondii

Presumptive involvement of methyl ricinoleate β-oxidation in the production of γ-decalactone by the yeastPichia guilliermondii

Production of γ-decalactone by yeasts from fatty acids has been reported but little is known about the mechanisms involved in this process. This paper provides information about the mechanisms involved in the production of γ-decalactone by Pichia guilliermondii in the presence of a fatty acid methyl ester. Culturing of P. guilliermondii in media containing methyl ricinoleate (12(R)-hydroxy-9(Z)-octadecenoic acid) revealed a coordinated induction of β-oxidation activities and γ-decalactone production. However, no γ-decalactone synthesis was noted when methyl ricinoleate was changed into methyloleate or methyl linoleate, even though these fatty acid methyl esters are able to induce β-oxidation activities in P. guilliermondii. These observations led us to conclude that methyl ricinoleate is an inducer of β-oxidation and is probably the substrate for γ-decalactone production. The fatty acid ester β-oxidation should be involved, at least in part, in this production.

The Production of γ-Decalactone by Fermentation of Castor Oil

A microbial process for the production of optically-active γ-decalactone from the ricinoleic acid present as triglycerides in castor oil has been developed, γ-decalactone (γDL) is a component of some fruit flavours, being an important organoleptic component of peach flavours. Screening showed two red yeast microorganisms, Rhodotorula glutinis and Sporobolomyces odortts to be especially suitable for this biotransformation. The process involves lipase-mediated hydrolysis of the castor oil to give free ricinoleic acid, uptake of the acid by the cells and aerobic fermentation to achieve abbreviated β-oxidation of the ricinoleic acid (12-hydroxyoleic acid) into 4-hydroxydecanoic acid (4HDA), lactonisation of the acid into γ-DL, followed by solvent extraction and distillation. γ-DL broth concentrations of 0.5-1.2g · 1-t were obtained after 3-5 days from fermentation media containing 10 g · 1-1 castor oil, representing an 8.3-20.0% theoretical yield. Intermediates detected were consistent with the operation of the β-oxidation pathway. Appreciable amounts of novel metabolites identified as cis and trans isomers of a tetrahydrofuran (C10) were also produced. Their formation from 4HDA appeared to be non-enzymic and was favoured by anaerobic conditions. Yields of γ-DL were inversely proportional to the concentration of castor oil present in the medium, indicating that substrate inhibition takes place. The highest yields of γ-DL were obtained when castor oil was present from the beginning of the fermentation, rather than when added once the fermentation had become established, demonstrating that the β-oxidation pathway and/or transport system require continual induction. Significant amounts of γ-DL were not produced from other fatty acids, including ricinelaidic acid, the trans isomer of ricinoleic acid. γ-DL formation was dramatically inhibited by antibiotic inhibitors of oxidative phosphorylation, indicating the importance of intact β-oxidation pathways, whereas inhibitors of protein synthesis and cell-wall synthesis had much less marked effects. Selective extraction of 4HDA from the fermentation broths, and of γDL from broth lactonised by heating at low pH, could be achieved by adsorption to Amberlite XAD-1 and XAD-7 resins respectively. Some product could be recovered from the exit gases of the fermenter by passing through propylene glycol traps. This pathway is unusual in that it is a rare example of the truncated β-oxidation of a fatty acid by microorganisms. This effect probably occurs because of partial inhibition of one or more enzymes of the β-oxidation pathway by the C10 hydroxylated fatty acid intermediate(s) allowing intracellular accumulation of the 4HDA, followed by leakage out of the cell; although further metabolism of this C10 intermediate does take place slowly.

Influence of Water Potential on γ-Decalactone Production by the Yeast Sporidiobolus salmonicolor

The influence of water potential on gamma-decalactone production by the yeast Sporidiobolus salmonicolor cultivated in a liquid medium was evaluated by gas-chromatographic analysis. Modifications in water potential led to a number of variations in the aroma production. Maximum extracellular production occurred at water activity (a(w)) with a value of 0.99. Further analyses revealed an important phenomenon of cellular accumulation of aroma for a(w) values between 0.97 and 0.99.