The dynamics of two wild type strains of Saccharomyces cerevisiae (BY4741 and EGY48) that vary in the ability to produce sterols were compared in batch cultures under different aeration conditions. Poor supply of oxygen enhanced selectivity of the bioprocess in favor of squalene formation. Optimization of inoculum size and fermentation time arranged according to a central composite statistical design revealed significant differences between the strains in terms of yield and productivity. Experimental verification showed that an optimized bioprocess under semianaerobic conditions is competitive with regard to those reported in the literature. Maximum squalene yield and productivity were, respectively, 2967.6 +/- 118.7 microg/L of culture medium and 104 +/- 4.2 microg/Lh for BY4741 and 3129 +/- 109.5 microg/L of culture medium and 155.9 +/- 5.5 microg/Lh for EGY48. The prospect of developing high-purity squalene preparations that meet food safety regulation demands is expected to attract the interest of the food industry.

Interest is increasing in establishing renewable sources for squalene, a functional lipid, as the conventional ones are limited. In the present study, squalene production was achieved in a wild-type laboratory Saccharomyces cerevisiae strain by two safe chemical means using terbinafine (0.05-0.55 mM) and methyl jasmonate (MJ) (0-1.00 mM). Bioprocess kinetics optimized by response surface methodology and monitored by high-performance liquid chromatography revealed a clear dependence of growth and squalene content (SQC) and yield (SQY) on the above regulators. Maximum SQC (10.02±0.53 mg/g dry biomass) and SQY (20.70±1.00 mg/L) were achieved using 0.442 mM terbinafine plus 0.044 mM MJ after 28 h and 0.300 mM terbinafine after 30 h, respectively. A 10-fold increase in SQY was achieved in comparison to that in the absence of regulator. The ruggedness of optimum conditions for SQY was verified for five industrial strains. The cellular lipid fraction (∼12% of dry biomass) was rich in squalene (12-13%). Results are encouraging toward bioprocess scale up.

Squalene uses extend from cosmetics to the medical and nutraceutical sectors. International concern for the protection of the deep sea sharks, the major source for this hydrocarbon so far, has engendered research interest in other directions (plant kingdom, microbes). Biotechnology offers an alternative approach with the potential of safety requirements and high-yield. Saccharomyces cerevisiae appears to be a promising choice for food and nutraceutical applications.

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Squalene (SQ), a precursor of sterols and terpenoids is a functional lipid of high importance in the food and pharmaceutical sectors. SQ oxidation studies are rather limited compared with those for other olefins. The aim of the present study was to monitor the formation of SQ oxidation products under different conditions (temperature, air supply), to characterise the most abundant of them by spectroscopic techniques and then examine their pro-oxidant activity in a model lipid substrate. Squalane (SQA), the saturated analogue of SQ, was used as a reference compound. FT-MIR analysis indicated the presence of alcohols, epoxides, aldehydes, and ketones. GC-MS was used to characterise SQ primary oxidation and scission products. The presence of epoxides was further confirmed by means of 1 H NMR and 13 C NMR spectroscopy. It could be argued that SQ stability is due to its stereochemistry and specifically to the presence of methyl groups next to the double bonds. The pro-oxidant activity of SQ oxidation products was evident at 62 and 40°C and suppressed only in the presence of primary antioxidants, not of SQ. The present work adds to the characterisation of SQ oxidised products. To our knowledge their pro-oxidant activity has never been examined before.Practical applications: Characterisation of squalene oxidation products and assessment of their activity as pro-oxidants present both scientific interest regarding the kinetics and product identity as well as a practical impact in case this bioactive lipid is provided for consumption as a functional product. In the past, cholesterol oxidation products and more recently phytosterol ones attracted the interest of researchers, who studied the stability of the respective parent compounds for food safety reasons.

The dynamics of industrial types of glycerol as a supplementary carbon source to glucose for beta-carotene production by Blakeslea trispora was investigated in batch cultures. The growth kinetics, cellular lipid accumulation-degradation, substrate assimilation, and beta-carotene production were clearly dependent on the level of addition of pure glycerol. The highest beta-carotene production (15.0 mg/g of dry biomass) was obtained at an initial glycerol concentration of 60.0 g/L. Substitution of pure glycerol by the nonpurified soap byproduct did not inhibit cell growth. Conversely, partial purification of the biodiesel byproduct by removing methanol and fatty acids was unavoidable for cell growth. Both types of industrial glycerol stimulated beta-carotene synthesis more than 10 (soap byproduct) and 8 times (biodiesel byproduct) compared to control medium. The maximum beta-carotene contents were 10 and 8 mg/g of dry biomass, respectively, and its relative content in the carotenoid fraction was 86-88%.

The present work is a systematic approach for valorization of wine lees regarding the recovery of squalene, a bioactive lipid. Such a study is presented for the first time in literature. Separate examination of squalene content in "light" and "heavy" lees from different vinification processes by RP-HPLC demonstrated that these waste streams can be used as a source for this lipid, despite variations due to technological or genetic effects. Next, ultrasound assisted extraction of squalene from the "industrial waste" (the mixture of wine lees generated from different wines) using n-hexane was optimized with the aid of response surface methodology (independent variables: sonication duration and duty cycles). Autolysis was monitored through optical microscopy. Squalene yield (0.6 ± 0.08 g SQ/kg dry lees) was comparable to that of recently examined potential sources (0.2-0.35 g SQ/kg dry olive pomace and 0.06 g SQ/kg olive leaves).