Cellic ctec2 cellulase

Poplar wood chips were provided by Shan Dong Sun Paper Industry Joint Stock Co., Ltd. (Shandong, China), they were milled into particles with a size range of 40–60 mesh; p-TsOH and some chromatographic grade organic solvents were supplied by Macklin biochemical Co., Ltd. (Shanghai, China); FA solution (37–40%, wt.%) and H₂SO₄ (95–98%, wt.%) were purchased from Yantai Far Eastern Fine Chemical CO., Ltd. (Shandong, China); 1,4-dioxane was obtained from Kemiou Chemical Reagent CO., Ltd. (Tianjin, China); DMSO-d₆ was provided by Cambridge Isotope Laboratories, Inc. (MA, United States). Cellulase (Cellic® CTec2) was supplied by Novozymes (Beijing, China).

Sugarcane bagasse was obtained from Guangxi Province, China. It was dried to constant weight at 60 °C and then stored in a polyethylene plastic container. The main components of sugarcane bagasse were as follows: 39.4 % glucan, 27.5 % xylan, and 26.0 % lignin. Industrial glycerol was of commercial grade (95 % purity), purchased from a chemical plant in Jiangsu Province, China. It was diluted to 70 % for use in the experiment. Cellulase Cellic CTec2 (150 FPU/g) was a generous gift from Novozymes (China) Investment Co. Ltd.

Cellobiose, glucose, xylose, arabinose, acetoin, acetic acid, lactic acid, and other chemicals used were of analytical grade. Cellulase Cellic CTec2 (Novozymes, Tianjin, China) was used for enzymatic saccharification of corncob.

Bamboo residues were provided by a bamboo processing factory in Sichuan, China. The air-dried bamboo powders were ground into particles (20–80 mesh) for subsequent experiments. The contents of glucan, xylan, and lignin in bamboo residues were 40.1% ± 0.2%, 22.0% ± 0.4%, and 27.2% ± 0.2%, respectively, which were analyzed according to the procedure developed by the US National Renewable Energy Laboratory (NREL). Cellulase (Cellic CTec2) was provided by Novozymes NA, Franklinton, USA with a filter paper activity of 250 FPU/ml.

The SCB and molasses used in this study were collected in the month of December from Maoyuan Sugar Co., Ltd., in Shaoguan, China. SCB was first milled into < 1 mm lengths with a MiniMill (MF10, IKA, Germany), sealed in a sealing bag, and stored at room temperature. The molasses sample was kept at 4 °C in a refrigerator for further utilization. The chemical compositions of untreated and pretreated SCB were determined using the method provided by the NREL [46 ]. The fermentation sugars in molasses were quantified by a high-performance liquid chromatography system (HPLC, Shimadzu, Japan) equipped with a refractive index detector (RID) and a cation-exchange column (SUGAR KS-801; 300 mm * 8.0 mm; Shodex™, Japan). The compositions of SCB and molasses are shown in Table 1.
Cellulase (Cellic CTec2) was obtained from Novozymes (Bagswade, Denmark), with an enzyme activity of 164 FPU/mL according to the methods of NREL [47 ].
The microorganism strain Saccharomyces cerevisiae was provided by Angel Yeast Co., Ltd. (Yichang, China).

Before the extraction, Fucus distichus subsp. evanescens brown algae was washed, lyophilized and grounded into a powder. The enzymatic extraction was performed as described in [21 (link)]. Briefly, dried seaweed was treated with Cellic^®CTec2 cellulase (Novozymes, Bagsværd, Denmark) and alginate lyase SALy from Sphingomonas sp. to break down non-fucoidan polysaccharide components of the cell and release fucoidan. High molecular weight alginate was removed by precipitating with CaCl₂. Fucoidans were further precipitated using ethanol. The crude fucoidan product was lyophilized for bioactivity studies (FE_crude). The aqueous solution of the crude fucoidan was further purified and fractionated using ion-exchange chromatography, obtaining three fractions (FE_F1, F2, F3) as previously described in [21 (link)]. Subsequently, the fractions were filtered through a 10 kDa membrane and lyophilized.