Extraction and Emulsification of Carotenoids from Carrot Pomaces Using Oleic Acid.

This study aimed to use oleic acid-based ultrasonic-assisted extraction (UAE) to recover carotenoids from carrot pomace and emulsify the enriched-carotenoid oleic acid using spontaneous and ultrasonic-assisted emulsification. The extraction performance of oleic acid was compared with traditional organic solvents, including hexane, acetone, and ethyl acetate. The one-factor experiments were employed to examine the impact of UAE conditions, including liquid-to-solid ratios, temperature, ultrasonic power, and time, on the extraction yield of carotenoids and to find the conditional ranges for the optimization process. The response surface methodology was employed to optimize the UAE process. The second-order extraction kinetic model was used to find the mechanism of oleic acid-based UAE. After that, the enriched-carotenoid oleic acid obtained at the optimal conditions of UAE was used to fabricate nanoemulsions using spontaneous emulsification (SE), ultrasonic-assisted emulsification (UE), and SE-UE. The effect of SE and UE conditions on the turbidity of nanoemulsion was determined. Then, the physiochemical attributes of the nanoemulsion from SE, UE, and spontaneous ultrasonic-assisted emulsification (SE-UE) were determined using the dynamic light scattering method. The extraction yield of carotenoids from carrot pomace by using sonication was the highest. The adjusted optimal conditions were 39 mL/g of LSR, 50 °C, 12.5 min, and 350 W of ultrasonic power. Under optimal conditions, the carotenoid content attained was approximately 163.43 ± 1.83 µg/g, with the anticipated value (166 µg/g). The particle sizes of nanoemulsion fabricated at the proper conditions of SE, UE, and SE-UE were 31.2 ± 0.83, 33.8 ± 0.52, and 109.7 ± 8.24 nm, respectively. The results showed that SE and UE are suitable methods for fabricating nanoemulsions. The research provided a green approach for extracting and emulsifying carotenoids from carrot pomace.

Biohythane production via single-stage fermentation using gel-entrapped anaerobic microorganisms: Effect of hydraulic retention time.

Research of single-stage anaerobic biohythane production is still in an infant stage. A single-stage dark fermentation system using separately-entrapped H2- and CH4-producing microbes was operated to produce biohythane at hydraulic retention times (HRTs) of 48, 36, 24, 12 and 6 h. Peak biohythane production was obtained at HRT 12 h with H2 and CH4 production rates of 3.16 and 4.25 L/L-d, respectively. At steady-state conditions, H2 content in biohythane and COD removal efficiency were in ranges of 7.3-84.6 % and 70.4-77.9%, respectively. During the fermentation, the microbial community structure of the entrapped H2-producing microbes was HRT-independent whereas entrapped CH4-producing microbes changed at HRTs 12 and 6 h. Caproiciproducens and Methanobacterium were the dominant genera for producing H2 and CH4, respectively. The novelty of this work is to develop a single-stage biohythane production system using entrapped anaerobic microbes which requires fewer controls than two-stage systems.

Biohythane production via single-stage anaerobic fermentation using entrapped hydrogenic and methanogenic bacteria.

This study demonstrates the continuous biohythane production in a single-stage anaerobic digester using a biomass mixture of separately entrapped hydrogenic and methanogenic bacteria (H2- and CH4-producing bacteria, respectively). The entrapped hydrogenic/methanogenic bacteria biomass ratios of 1/4, 2/3, 3/2 and 4/1 were tested and shown to have a great effect on the single-stage biohythane production performance. At steady-states, the cultivations had biohythane production rates in the range of 381-480 mL/L-d, with H2 content in biohythane (HCH) varying from 1% to 75% (v/v) and chemical oxygen demand removal efficiencies (TCODre) of 57.6-81.9%. Biomass ratio 2/3 (weight ratio 1/1.5) resulted in peak biohythane production with H2 and CH4 production rates being 64.6 and 395 mL/L-d, respectively, HCH 15% and TCODre 74.4%. The novelty of this work is to show the potential of producing biohythane from an innovative single-stage dark fermentation system using entrapped hydrogenic and methanogenic bacteria.

A shift to 50°C provokes death in distinct ways for glucose- and oleate-grown cells of Yarrowia lipolytica.

Based on the observation that shocks provoked by heat or amphiphilic compounds present some similarities, this work aims at studying whether cells grown on oleate (amphiphilic pre-stress) acquire a tolerance to heat shock. In rich media, changing glucose for oleate significantly enhanced the cell resistance to the shock, however, cells grown on a minimal oleate medium lost their ability to grow on agar with the same kinetic than glucose-grown cells (more than 7-log decrease in 18 min compared with 3-log for oleate-grown cells). Despite this difference in kinetics, the sequence of events was similar for oleate-grown cells maintained at 50°C with a (1) loss of ability to form colonies at 27°C, (2) loss of membrane integrity and (3) lysis (observed only for some minimal-oleate-grown cells). Glucose-grown cells underwent different changes. Their membranes, which were less fluid, lost their integrity as well and cells were rapidly inactivated. But, surprisingly, their nuclear DNA was not stained by propidium iodide and other cationic fluorescent DNA-specific probes but became stainable by hydrophobic ones. Moreover, they underwent a dramatic increase in membrane viscosity. The evolution of lipid bodies during the heat shock depended also on the growth medium. In glucose-grown cells, they seemed to coalesce with the nuclear membrane whereas for oleate-grown cells, they coalesced together forming big droplets which could be released in the medium. In some rare cases of oleate-grown cells, lipid bodies were fragmented and occupied all the cell volume. These results show that heat triggers programmed cell death with uncommon hallmarks for glucose-grown cells and necrosis for methyl-oleate-grown cells.

Biochemistry of lactone formation in yeast and fungi and its utilisation for the production of flavour and fragrance compounds.

The consumers' demand for natural flavour and fragrances rises. To be natural, compounds have to result from the extraction of natural materials and/or to be transformed by natural means such as the use of enzymes or whole cells. Fungi are able to transform some fatty acids into lactones that can thus be natural. Although some parts of this subject have been reviewed several times, the present article proposes to review the different pathways utilised, the metabolic engineering strategies and some current concerns on the reactor application of the transformation including scaling up data. The main enzymatic steps are hydroxylation and ß-oxidation in the traditional way, and lactone desaturation or Baeyer-Villiger oxidation. Although the pathway to produce γ-decalactone is rather well known, metabolic engineering strategies may result in significant improvements in the productivity. For the production of other lactones, a key step is the hydroxylation of fatty acids. Beside the biotransformation, increasing the production of the various lactones requires from biotechnologists to solve two main problems which are the toxicity of lactones toward the producing cell and the aeration of the emulsified reactor as the biochemical pathway is very sensitive to the level of available oxygen. The strategies employed to resolve these problems will be presented.

New insights into the effect of medium-chain-length lactones on yeast membranes. Importance of the culture medium.

In hydrophobic compounds biotechnology, medium-chain-length metabolites often perturb cell activity. Their effect is usually studied in model conditions of growth in glucose media. Here, we study whether culture on lipids has an impact on the resistance of Yarrowia lipolytica to such compounds: Cells were cultured on glucose or oleate and submitted to gamma-dodecalactone. After a 60-min exposure to 3 g L(-1), about 80% of the glucose-grown cells (yeast extract peptone dextrose (YPD) cells) had lost their cultivability, 38% their membrane integrity, and 31% their reducing capacity as shown with propidium iodide and methylene blue, respectively. For oleate-grown cells, treatment at 6 g L(-1) did not alter cultivability despite some transient loss of membrane integrity from 3 g L(-1). It was shown with diphenylhexatriene and 1-(4-trimethylammoniumphenyl)-6-phenyl-1,3,5-hexatriene that oleate-grown cells had membranes more fluid and less sensitive to the lactone-induced fluidization. Analyses revealed also higher contents of ergosterol but, for YPD- and minimum-oleate-grown cells (YNBO cells), the addition of lactone provoked a decrease in the concentration of ergosterol in a way similar to the depletion by methyl-beta-cyclodextrin and an important membrane fluidization. Ergosterol depletion or incorporation increased or decreased, respectively, cell sensitivity to lactone. This study shows that the embedment of oleate moieties into membranes as well as higher concentrations of sterol play a role in the higher resistance to lactone of oleate-grown cells (YPO cells). Similar oleate-induced increase in resistance was also observed for Rhodotorula and Candida strains able to grow on oleate as the sole carbon source whereas Saccharomyces and Sporidiobolus cells were more sensitive after induction.