Polyhydroxyalkanoates (PHAs) are an important class of biodegradable polymers synthesized by a few bacteria under nutrient-limiting conditions. In this study, the lipase-catalysed degradation of PHA synthesized by Enterobacter sp. was monitored. For this, the lipase-encoding gene from Bacillus subtilis DI2 was PCR-amplified, cloned into a T vector system and sequenced. It was expressed in Escherichia coli DH5a cells, the recombinant enzyme was purified 24.25-fold, and its molecular weight was determined to be around 28 kDa. When PHA biodegradation studies were carried out with this enzyme, gel permeation chromatography showed 21.3 and 28.3 % molecular weight decrease and weight loss, respectively. Further, scanning electron micrographs revealed alterations in polymer surface morphology. Changes in molecular vibrations were noticed in the FTIR spectra. When the chemical shifts in NMR spectra were studied, a steep reduction in area under the peak at 1.57 ppm was observed. In the heating range of 30-930°C employed during thermogravimetry analysis, the degraded sample showed a total of 45.82 % weight loss, as against 18.89 % for the native sample. The melting temperature (T m ) of the polymer was also brought down from 126.22 to 118.18°C, as inferred from differential scanning calorimetry. Lipase-catalysed chain scission reactions could thus be used to generate low molecular weight functional biopolymers with wide-ranging pharmaceutical applications, such as in sustained drug release.

Steam explosion is a well-known process to pretreat lignocellulosic biomass in order to enhance sugar yields in enzymatic hydrolysis, but pretreatment conditions have to be optimized individually for each material. In this study, we investigated how the results of a pretreatment optimization procedure are influenced by the chosen reaction conditions in the enzymatic hydrolysis. Beechwood was pretreated by steam explosion and the resulting biomass was subjected to enzymatic hydrolysis at glucan loadings of 1% and 5% employing either washed solids or the whole pretreatment slurry. For enzymatic hydrolysis in both reaction modes at a glucan loading of 1%, the glucose yields markedly increased with increasing severity and with increasing pretreatment temperature at identical severities and maximal values were reached at a pretreatment temperature of 230 °C. However, the optimal severity was 5.0 for washed solids enzymatic hydrolysis, but only 4.75 for whole slurry enzymatic hydrolysis. When the glucan loading was increased to 5%, glucose yields hardly increased for pretreatment temperatures between 210 and 230 °C at a given severity, and a pretreatment temperature of 220 °C was sufficient under these conditions. Consequently, it is important to precisely choose the desired conditions of the enzymatic hydrolysis reaction, when aiming to optimize the pretreatment conditions for a certain biomass.

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Biomass pretreatment is a mandatory step for the biochemical conversion of lignocellulose to chemicals. During pretreatment, soluble compounds are released into the prehydrolyzate that inhibit the enzymatic hydrolysis step. In this work, we investigated how the reaction conditions in steam explosion pretreatment of beechwood (severity: 3.0–5.25; temperature: 160–230 °C) influence the resulting amounts of different inhibitors. Furthermore, we quantified the extent of enzyme inhibition during enzymatic hydrolysis of Avicel in the presence of the prehydrolyzates. The amounts of phenolics, HMF, acetic acid and formic acid increased with increasing pretreatment severities and maximal quantities of 21.6, 8.3, 43.7 and 10.9 mg/gbeechwood, respectively, were measured at the highest severity. In contrast, the furfural concentration peaked at a temperature of 200 °C and a severity of 4.75. The presence of the prehydrolyzates in enzymatic hydrolysis of Avicel lowered the glucose yields by 5–26%. Mainly, the amount of phenolics and xylose and xylooligomers contributed to the reduced yield. As the maximal amounts of these two inhibitors can be found at different conditions, a wide range of pretreatment severities led to severely inhibiting prehydrolyzates. This study may provide guidelines when choosing optimal pretreatment conditions for whole slurry enzymatic hydrolysis.

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