Ics 3000 hplc

HPAEC analysis was performed on a ICS-3000 HPLC (Dionex Corporation) using a CarboPac PA200 analytical column (3 × 150 mm) and a CarboPac PA200 guard column (3 × 30 mm) at 30 °C. Following injection of 25 μL of diluted samples, elution was performed at 0.4 mL∕ min using 0.1 M NaOH in the mobile phase with sodium acetate gradients. For aldonic acid separation, the acetate gradients were 0 mM for 1 min, increasing to 80 mM in 8 min, increasing to 300 mM in 2 min, keeping at 300 mM for 1 min, increasing to 600 mM for 2 min, keeping at 600 mM for 5 min, and re-equilibrating at 0 mM for 4 min. Elution was monitored using pulsed amperometric detection (PAD) and peaks were analyzed and quantified using the Chromeleon software package.

Carbohydrates were measured on a Dionex (Sunnyvale, CA, USA) HPLC (ICS-3000) system equipped with an autosampler, electrochemical detector, dual pumps, and anion exchange column (Dionex, CarboPac PA1). Deionized water at 1 ml/min was used as an eluent, and post-column addition of 0.2 M NaOH at a flow rate of 0.5 ml/min ensured optimization of baseline stability and detector sensitivity. After each analysis, the column was reconditioned with 0.25 M NaOH. Twenty microliters of each sample were injected after filtration through a 0.22-µm syringe filter (Restek Corp., Bellefonte, PA, USA). Samples were measured against standards consisting of arabinose, galactose, glucose, xylose, and mannose. In addition, fucose was used as an internal standard.
Acetic acid, furfural, and HMF were measured using refractive index detection on a Shimadzu Prominence LC. Separation of these compounds was achieved by an anion exchange column [REZEX RHM-Mono-saccharide H⁺ (8 %), Phenomenex, Inc., Torrance, CA, USA] with an isocratic mobile phase that of 5 mM H₂SO₄ at a flow rate of 0.6 ml/min. The column oven temperature was maintained at 63 °C. Twenty microliters of each sample were injected after being appropriately diluted in deionized water and filtered through a 0.22-µm syringe filter (Restek Corp., Bellefonte, PA, USA). Standards were prepared and used to quantify the unknown samples.

The carbohydrate and lignin contents of the raw and pretreated bagasses were determined as previously described [31 (link)]. Briefly, the carbohydrates in the substrates were acid hydrolyzed and the sugars were quantified with a Dionex (Sunnyvale, CA) HPLC (ICS-3000). Glucan content was determinate based on the released glucose, and xylan content on the released xylose, arabinose, and acetic acid. The acid insoluble lignin was gravimetrically analyzed and the acid soluble lignin was measured by reading the absorbance at 205 nm, using an extinction coefficient of 105 L g⁻¹ cm⁻¹ [52 ].

The concentration of monomeric sugars from chemical composition analyses and enzymatic hydrolysis was determined with a Dionex (Sunnyvale, CA) HPLC (ICS-3000) system equipped with an autosampler, dual pumps, an anion-exchange column (Dionex, CarboPac PA1), and an electrochemical detector (Dionex disposable gold electrode) [15 (link)]. DI water at 1 mL/min was used as an eluent, and postcolumn addition of 0.2 M NaOH at a flow rate of 0.5 mL/min ensured optimization of baseline stability and detector sensitivity. Acetic acid, furfural, HMF, and ethanol were measured using refractive index detection on a Shimadzu Prominence LC. Separation of these compounds was achieved by an anion-exchange column (Rezex RHM Monosaccharide H⁺ (8%), Phenomenex, Inc., Torrance, CA) with an isocratic mobile phase that consisted of 5 mM H₂SO₄ at a flow rate of 0.6 mL/min [15 (link)].