Effect of Inoculum Size and Age, and Sucrose Concentration on Cell Growth to Promote Metabolites Production in Cultured Taraxacum officinale (Weber) Cells.

Pentacyclic triterpenes, including lupeol, α- amyrin, and ß-amyrin, present a large range of biological activities including anti-inflammatory, anti-cancer, and gastroprotective properties. The phytochemistry of dandelion (Taraxacum officinale) tissues has been widely described. Plant biotechnology offers an alternative for secondary metabolite production and several active plant ingredients are already synthesized through in vitro cultures. This study aimed to establish a suitable protocol for cell growth and to determine the accumulation of α-amyrin and lupeol in cell suspension cultures of T. officinale under different culture conditions. To this end, inoculum density (0.2% to 8% (w/v)), inoculum age (2- to 10-week-old), and carbon source concentration (1%, 2.3%, 3.2%, and 5.5% (w/v)) were investigated. Hypocotyl explants of T. officinale were used for callus induction. Age, size, and sucrose concentrations were statistically significant in cell growth (fresh and dry weight), cell quality (aggregation, differentiation, viability), and triterpenes yield. The best conditions for establishing a suspension culture were achieved by using a 6-week-old callus at 4% (w/v) and 1% (w/v) of sucrose concentration. Results indicate that 0.04 (±0.02) α-amyrin and 0.03 (±0.01) mg/g lupeol can be obtained in suspension culture under these starting conditions at the 8th week of culture. The results of the present study provide a backdrop for future studies in which an elicitor could be incorporated to increase the large-scale production of α-amyrin and lupeol from T. officinale.

Effect of growth conditions on cell wall composition and cadmium adsorption in Chlorella vulgaris: A new approach to biosorption research.

Biosorption of toxic metals in microalgae is a process relying on the presence of cell wall reactive groups acting as binding sites. This work studied the effect of culture conditions on the outer cell wall composition of C. vulgaris and cadmium biosorption. The experiments were conducted in continuous culture under light and nitrogen limitation at two growth rates (0.4 and 0.2 d-1). Functional groups were profiled using ATR-FTIR spectrometry, and total cadmium biosorption was assayed. Significant differences in composition were attested the most salient being the absence of carboxyl groups in the light deprived states and a larger number of carbohydrates and amino groups in the nitrogen deprived cultures, particularly amino groups from deacetylated D-glucosamine polysaccharides. Higher biosorption was obtained with the nitrogen-restricted biomass, reaching a maximum of 11.9 mgCd/gbiomass, as compared to a minimum of 8.0 mgCd/gbiomass achieved in the light-restricted states. The increased biosorption exhibited by nitrogen-restricted strains was attributed to the deacetylated amino groups that have enhanced cation affinity. This work has shown that the characteristics of the outer cell wall can be engineered by culture conditions to improve biosorption, providing a new approach that opens up new research frontiers for the biosorption of hazardous metals.

Kinetics and modeling of cell growth for potential anthocyanin induction in cultures of Taraxacum officinale G.H. Weber ex Wiggers (Dandelion) in vitro

Background: Taraxacum officinale G.H. Weber ex Wiggers is a wild plant used in folk medicine to treat several diseases owing to bioactive secondary metabolites present in its tissue. The accumulation of such molecules in plant cells can occur as a response against abiotic stress, but these metabolites are often deposited in low concentrations. For this reason, the use of a biotechnological approach to improve the yields of technologically interesting bioactive compounds such as anthocyanins is a compelling option. This work focuses on investigating the potential of in vitro T. officinale cultures as an anthocyanin source. Results: To demonstrate the suitability of anthocyanin induction and accumulation in calluses under specific conditions, anthocyanin was induced in the T. officinale callus. A specific medium of 5.5% sucrose supplemented with 6-benzylaminopurine /1-naphthaleneacetic acid in a 10:1 ratio was used to produce an anthocyanin yield of 1.23 mg g-1 fw. An in vitro dandelion callus line was established from this experiment. Five mathematical models were then used to objectively and predictably explain the growth of anthocyanin-induced calluses from T. officinale. Of these models, the Richards model offered the most suitable representation of anthocyanin callus growth in a solid medium and permitted the calculation of the corresponding kinetic parameters. Conclusions: The findings demonstrate the potential of an in vitro anthocyanin-induced callus line from T. officinale as an industrial anthocyanin source.

Influence of hydraulic retention time on the psychrophilic hydrolysis/acidogenesis of proteins.

The influence of the hydraulic retention time (HRT) on the anaerobic hydrolysis of complex substrates has been studied under psychrophilic conditions. For this purpose, a continuous stirred tank reactor was operated at 15 °C and neutral pH and gelatin was considered as a model protein. Three HRTs have been tested: 12, 21 and 36 h. Gelatin hydrolysis was greatly dependent on HRT, increasing from 40% at 12 h-HRT to a maximum of 65% at 36 h-HRT. Molecular size distribution analyses of the effluent showed that hydrolysation of compounds larger than 10 kDa was poor at 12 h-HRT, whereas the fraction of 1-10 kDa was completely transformed into compounds smaller than 1 kDa. Higher HRT (36 h) improved the degradation of the recalcitrant fraction (>10 kDa), obtaining an effluent with around 95% of soluble molecules (<1 kDa). In that way, the use of membrane bioreactors for the treatment of this type of macromolecules could improve the degradation efficiencies by enabling to increase the residence time of the non-hydrolyzed molecules, with what would be possible to achieve higher organic loading rate operation.

The enhancement of antioxidant compounds extracted from Thymus vulgaris using enzymes and the effect of extracting solvent.

We evaluate the total phenolic compounds (TPC) content and the antioxidant activity (AA) of extracts obtained from ground fresh thyme (FT) and depleted thyme (DT), a by-product of the process of essential oil extraction. In addition, enzymatic treatments were evaluated to improve the extraction yields of polyphenolic compounds from thyme. Extractions were performed using several solvents as methanol, ethanol, and water. Enzymes were applied prior to extraction or during the extraction process. The best results were obtained using a mixture of methanol and water, resulting in 2790 and 220 mg Gallic acid equivalent (GAE)/L of TPC for FT and DT, respectively. A similar result was observed for AA. With regard to enzymatic treatment, application of Grindamyl CA 150 enzyme as a pre-treatment resulted in the production of an extract from DT with 614 mg TE (trolox equivalent)/L of AA, 70% more than the control, and an AA of 621 mg TE/L (74% more than the control sample) was obtained using Grindamyl CA 150 during the extraction process. These results suggest that enzymatic treatment is an interesting alternative for producing antioxidant extracts from DT.

Improved sanitary landfill design using recirculation of anaerobically treated leachates: generation of advanced design criteria

In Latin America, the most accepted disposal systems for Municipal Solid Waste (MSW) are landfills, which nowadays have low rates of stabilization. The objective of this study was to develop design criteria for sanitary landfills which lead to a reduction in the stabilization times of MSW, based on experiment results obtain from a pre-pilot scale operation of two sanitary landfills (0.5 Ton), one with recirculation of leachates treated in an anaerobic digester and the other with recirculation of untreated leachates. This was complemented by another pilot scale sanitary landfill (1440 Ton) with recirculation of leaches treated in an anaerobic filter, and additionally by a computer simulation of leachate generation through the water balance of a theoretical cell of MSW (850 Ton), in which the initial humidity of the MSW and the type of final cover were evaluated. The results obtained on the pre-pilot scale indicate that recirculation of anaerobically treated leachates, when compared to the recirculation of untreated leachates, increased the rate of MSW stabilization, projecting a stabilization time reduction of 72 percent. In the pilot sanitary landfill an increase in the settling rate of around 200 percent was observed when operating with the recirculation of anaerobically treated leachates as opposed to operation without recirculation. The water balance carried out on the theoretical cell of MSW demonstrated the importance to leachate generation of both the initial landfill water saturation and the type of final barrier. From these results it may be stated that it is important for landfill design to maintain waste humidity close to its field capacity along with an adequate leachate application rate.