Overcoming factors limiting high-solids fermentation of lignocellulosic biomass to ethanol.

Simultaneous saccharification and fermentation (SSF) of solid biomass can reduce the complexity and improve the economics of lignocellulosic ethanol production by consolidating process steps and reducing end-product inhibition of enzymes compared with separate hydrolysis and fermentation (SHF). However, a long-standing limitation of SSF has been too low ethanol yields at the high-solids loading of biomass needed during fermentation to realize sufficiently high ethanol titers favorable for more economical ethanol recovery. Here, we illustrate how competing factors that limit ethanol yields during high-solids fermentations are overcome by integrating newly developed cosolvent-enhanced lignocellulosic fractionation (CELF) pretreatment with SSF. First, fed-batch glucose fermentations by Saccharomyces cerevisiae D5A revealed that this strain, which has been favored for SSF, can produce ethanol at titers of up to 86 g⋅L-1 Then, optimizing SSF of CELF-pretreated corn stover achieved unprecedented ethanol titers of 79.2, 81.3, and 85.6 g⋅L-1 in batch shake flask, corresponding to ethanol yields of 90.5%, 86.1%, and 80.8% at solids loadings of 20.0 wt %, 21.5 wt %, and 23.0 wt %, respectively. Ethanol yields remained at over 90% despite reducing enzyme loading to only 10 mg protein⋅g glucan-1 [∼6.5 filter paper units (FPU)], revealing that the enduring factors limiting further ethanol production were reduced cell viability and glucose uptake by D5A and not loss of enzyme activity or mixing issues, thereby demonstrating an SSF-based process that was limited by a strain's metabolic capabilities and tolerance to ethanol.

Nanophase hydroxyapatite and poly(lactide-co-glycolide) composites promote human mesenchymal stem cell adhesion and osteogenic differentiation in vitro.

Human mesenchymal stem cells (hMSCs) typically range in size from 10 to 50 µm and proteins that mediate hMSC adhesion and differentiation usually have a size of a few nanometers. Nanomaterials with a feature size smaller than 100 nm have demonstrated the unique capability of promoting osteoblast (bone forming cell) adhesion and long-term functions, leading to more effective bone tissue regeneration. For new bone deposition, MSCs have to be recruited to the injury or disease sites and then differentiate into osteoblasts. Therefore, designing novel nanomaterials that are capable of attracting MSCs and directing their differentiation is of great interest to many clinical applications. This in vitro study investigated the effects of nanophase hydroxyapatite (nano-HA), nano-HA/poly(lactide-co-glycolide) (PLGA) composites and a bone morphogenetic protein (BMP-7) derived short peptide on osteogenic differentiation of hMSCs. The short peptide was loaded by physical adsorption to nano-HA or by dispersion in nanocomposites and in PLGA to determine their effects on hMSC adhesion and differentiation. The results showed that the nano-HA/PLGA composites promoted hMSC adhesion as compared to the PLGA controls. Moreover, nano-HA/PLGA composites promoted osteogenic differentiation of hMSCs to a similar extent with or without the presence of osteogenic factors in the media. In the MSC growth media without the osteogenic factors, the nanocomposites supported greater calcium-containing bone mineral deposition by hMSC than the BMP-derived short peptide alone. The nanocomposites provided promising alternatives in controlling the adhesion and differentiation of hMSCs without osteogenic factors from the culture media, and, thus, should be further studied for clinical translation and the development of novel nanocomposite-guided stem cell therapies.

Co-solvent pretreatment reduces costly enzyme requirements for high sugar and ethanol yields from lignocellulosic biomass.

We introduce a new pretreatment called co-solvent-enhanced lignocellulosic fractionation (CELF) to reduce enzyme costs dramatically for high sugar yields from hemicellulose and cellulose, which is essential for the low-cost conversion of biomass to fuels. CELF employs THF miscible with aqueous dilute acid to obtain up to 95 % theoretical yield of glucose, xylose, and arabinose from corn stover even if coupled with enzymatic hydrolysis at only 2 mgenzyme gglucan (-1) . The unusually high saccharification with such low enzyme loadings can be attributed to a very high lignin removal, which is supported by compositional analysis, fractal kinetic modeling, and SEM imaging. Subsequently, nearly pure lignin product can be precipitated by the evaporation of volatile THF for recovery and recycling. Simultaneous saccharification and fermentation of CELF-pretreated solids with low enzyme loadings and Saccharomyces cerevisiae produced twice as much ethanol as that from dilute-acid-pretreated solids if both were optimized for corn stover.

The effects of poly(3,4-ethylenedioxythiophene) coating on magnesium degradation and cytocompatibility with human embryonic stem cells for potential neural applications.

Magnesium (Mg) is a promising conductive metallic biomaterial due to its desirable mechanical properties for load bearing and biodegradability in human body. Controlling the rapid degradation of Mg in physiological environment continues to be the key challenge toward clinical translation. In this study, we investigated the effects of conductive poly(3,4-ethylenedioxythiophene) (PEDOT) coating on the degradation behavior of Mg substrates and their cytocompatibility. Human embryonic stem cells (hESCs) were used as the in vitro model system to study cellular responses to Mg degradation because they are sensitive and can potentially differentiate into many cell types of interest (e.g., neurons) for regenerative medicine. The PEDOT was deposited on Mg substrates using electrochemical deposition. The greater number of cyclic voltammetry (CV) cycles yielded thicker PEDOT coatings on Mg substrates. Specifically, the coatings produced by 2, 5, and 10 CV cycles (denoted as 2×-PEDOT-Mg, 5×-PEDOT-Mg, and 10×-PEDOT-Mg) had an average thickness of 31, 63, and 78 µm, respectively. Compared with non-coated Mg samples, all PEDOT coated Mg samples showed slower degradation rates, as indicated by Tafel test results and Mg ion concentrations in the post-culture media. The 5×-PEDOT-Mg showed the best coating adhesion and slowest Mg degradation among the tested samples. Moreover, hESCs survived for the longest period when cultured with the 5×-PEDOT-Mg samples compared with the non-coated Mg and 2×-PEDOT-Mg. Overall, the results of this study showed promise in using PEDOT coating on biodegradable Mg-based implants for potential neural recording, stimulation and tissue engineering applications, thus encouraging further research.

Effects of magnesium on growth and proliferation of human embryonic stem cells.

The effects of magnesium on the growth and proliferation of human embryonic stem cells (hESCs) was explored to advance magnesium as an implant biomaterial. When magnesium ions from magnesium salt were added to the culture media at 10, 100, 250, 500, 750, and 1000 ppm (0.4, 4, 10, 20, 30, 40 mM) the rate of increase in viable cell coverage over time was higher for the larger doses of magnesium salt. Thus, the addition of magnesium ions exerted a positive effect on viable cell coverage. When hESCs were cultured with pure magnesium metal strips through transwell inserts, the cells underwent an initial increase in viable cell coverage, followed by rapid cell death within the first 24 hours. This initial increase in viable cell coverage corresponded to the colonies dispersing and losing their tightly packed morphologies. The cell death may be attributed to an increased alkalinity in the culture media incubated with the magnesium metal strips. In conclusion, since the degradation of magnesium results in both magnesium ions and OH- ions (an increase of pH), controlling the degradation of magnesium to obtain the perfect balance of ions is critical for advancing magnesium as an implant biomaterial.