BackgroundWinter annual crops such as winter rye (Secale cereale L) can produce biomass feedstock on seasonally fallow land that continues to provide high-value food and feed from summer annuals such as corn and soybeans. As energy double crops, winter grasses are likely to be harvested while still immature and thus structurally different from the fully senesced plant material typically used for biofuels. This study investigates the dynamic trends in biomass yield, composition, and biological solubilization over the course of a spring harvest season.ResultsThe water soluble fraction decreased with increasing maturity while total carbohydrate content stayed roughly constant at about 65%. The protein mass fraction decreased with increasing maturity, but was counterbalanced by increasing harvest yield resulting in similar total protein across harvest dates. Winter rye was ground and autoclaved then fermented at 15 g/L total solids by either (1) Clostridium thermocellum or (2) simultaneous saccharification and cofermentation (SSCF) using commercial cellulases (CTec2 and HTec2) and a xylose-fermenting Saccharomyces cerevisiae strain. Solubilization of total carbohydrate dropped significantly as winter rye matured for both C. thermocellum (from approximately 80% to approximately 50%) and SSCF (from approximately 60% to approximately 30%). C. thermocellum achieved total solubilization 33% higher than that of SSCF for the earliest harvest date and 50% higher for the latest harvest date. Potential revenue from protein and bioethanol was stable over a range of different harvest dates, with most of the revenue due to ethanol. In a crop rotation with soybean, recovery of the soluble protein from winter rye could increase per hectare protein production by 20 to 35%.ConclusionsDouble-cropping winter rye can produce significant biomass for biofuel production and feed protein as coproduct without competing with the main summer crop. During a 24-day harvest window, the total carbohydrate content remained relatively constant while the early-harvest yielded much higher carbohydrate solubilization for both C. thermocellum fermentation and SSCF. C. thermocellum fermentation achieved higher carbohydrate solubilization than SSCF across all growth stages tested. Although winter rye’s yield, composition, and biological reactivity change rapidly in the spring, it offers a substantial and stable income across the harvest season and thus flexibility for the farmer.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-015-0225-z) contains supplementary material, which is available to authorized users.

Accurate analysis of degradability of silage neutral detergent fiber (NDF) is important for diet formulation and to predict lactational performance of dairy cows. In this study, 5 corn silage hybrids ensiled for 0 (unfermented), 30, 60, 120, and 150 d were used to determine the effects of ensiling time on silage neutral detergent fiber degradability (NDFD) and to assess the relationships between near-infrared reflectance spectroscopy (NIR) NDF-related analyses and in situ NDFD variables. In addition, the relationships between dietary concentration of indigestible NDF, 288-h incubation (iNDF288), or undegraded NDF, 240-h incubation (uNDF240), and in vivo total-tract apparent organic matter and NDF digestibility were studied in total mixed ration samples from 16 experiments with lactating dairy cows. Ensiling time had no effect on silage NDF concentration; however, the ratio of acid detergent fiber ÷ NDF increased, and estimated hemicellulose concentration decreased quadratically with ensiling time. Also, concentration of NDF-bound protein decreased, and that of lignin increased linearly with ensiling time. These changes in silage fiber composition resulted in a linear decrease in in situ effective degradability of silage NDF with increasing ensiling time. The indigestible fraction of NDF and concentration of structural carbohydrates were not affected by ensiling time. Correlations of in situ NDFD variables with laboratory NIR NDFD analyses were weak to moderate. The relationship of corn silage uNDF240 with lignin concentration or 30-h NDFD (all NIR analyses) was remarkably good (R 2 = 0.73 and 0.88, respectively). The relationship between in situ iNDF288 concentration (but not uNDF240) and in vivo total-tract apparent digestibility of dietary organic matter and NDF was good (R 2 = 0.72 and 0.80, respectively). In conclusion, in situ degradability of silage NDF linearly decreased from 0 to 150 d ensiling time, primarily caused by a decrease in concentrations of hemicellulose and NDF-bound protein. In situ NDF degradability measurements and common laboratory NIR NDF-related analyses were generally poorly correlated. We found a good relationship between in vivo NDF digestibility and dietary concentration of iNDF288 determined in situ, but the relationship with uNDF240 was poor.

Commercial scale production of biofuels from lignocellulosic feed stocks has been hampered by the resistance of plant cell walls to enzymatic conversion, primarily owing to lignin. This study investigated whether DypB, the lignin-degrading peroxidase from Rodococcus jostii, depolymerizes lignin and reduces recalcitrance in transgenic tobacco (Nicotiana benthamiana). The protein was targeted to the cytosol or the ER using ER-targeting and retention signal peptides. For each construct, five independent transgenic lines were characterized phenotypically and genotypically. Our findings reveal that expression of DypB in the cytosol and ER does not affect plant development. ER-targeting increased protein accumulation, and extracts from transgenic leaves showed higher activity on classic peroxidase substrates than the control. Intriguingly, in situ DypB activation and subsequent saccharification released nearly 200% more fermentable sugars from transgenic lines than controls, which were not explained by variation in initial structural and non-structural carbohydrates and lignin content. Pyrolysis-GC-MS analysis showed more reduction in the level of lignin associated pyrolysates in the transgenic lines than the control primarily when the enzyme is activated prior to pyrolysis, consistent with increased lignin degradation and improved saccharification. The findings reveal for the first time that accumulation and in situ activation of a peroxidase improves biomass digestibility.

To understand molecular effects of ultrasound on protein gels (cross-linked, hydrated macromolecular systems of immeasurably high macroviscosity, but low microviscosity), the thick fraction of hen albumen was sonicated. The immeasurably high viscosity of the intact thick fraction decreased to 2.5-4.0 mPa·s (depending on the sample) after a 12 min sonication (0.14 mM of radicals were produced and 19 J g(-1) of thermal energy absorbed) indicating that the 3D protein network was degraded. SDS-PAGE analysis indicated the breaking of intermolecular S-S bridges holding together the protein network rather than the primary structure of constituent proteins. Despite the relatively large concentration of OH radical produced in the sonication time range applied, no protein cross-linking was observed which can be attributed to the high degree of protein glycosylation and protein immobility. Differential scanning calorimetry (DSC) showed that both the amount of bound water and the enthalpy of denaturation of the constituent proteins are not affected by sonication, which is consistent with the SDS-PAGE results. A small increase in sample turbidity can be attributed to the small extent of thermal denaturation occurring in the vicinity of cavitation sites.

Crop-based bioethanol has raised concerns about competition with food and feed supplies, and technologies for second- and third-generation biofuels are still under development. Alternative feedstocks could fill this gap if they can be converted to biofuels using current sugar- or starch-to-ethanol technologies. The aim of this study was to enhance carbohydrate accumulation in transgenic Nicotiana benthamiana by simultaneously expressing the maize Corngrass1 miRNA (Cg1) and E. coli ADP-glucose pyrophosphorylase (glgC), both of which have been reported to enhance carbohydrate accumulation in planta. Our findings revealed that expression of Cg1 alone increased shoot branching, delayed flowering, reduced flower organ size, and induced loss of fertility. These changes were fully restored by coexpressing Escherichia coli glgC. The transcript level of miRNA156 target SQUAMOSA promoter binding-like (SPL) transcription factors was suppressed severely in Cg1-expressing lines as compared to the wild type. Expression of glgC alone or in combination with Cg1 enhanced biomass yield and total sugar content per plant, suggesting the potential of these genes in improving economically important biofuel feedstocks. A possible mechanism of the Cg1 phenotype is discussed. However, a more detailed study including genome-wide transcriptome and metabolic analysis is needed to determine the underlying genetic elements and pathways regulating the observed developmental and metabolic changes.