Vp series liquid chromatograph

To prepare the RJ water/media extracts, 2 g of RJ were either mixed with 2 ml double distilled water for the physicochemical analysis or with 2 ml of the respective medium for the bacterial growth assay. Samples were centrifuged twice for 10 min at 20,000g to ensure the removal of any solids. Glucose, fructose, and sucrose contents were determined by HPLC (Sesta, 2006) using a Shimadzu VP series liquid chromatograph equipped with a degasser, pump, controller, column oven and an auto injector (Shimadzu Scientific Instruments, Columbia, USA). Acetonitril/water 75% (v/v) was used as the mobile phase for the separation of sugars on an Alltima Amino 100Å 5 μm column (256 × 4.6 mm) at a flow rate of 1.3 ml/min, 30°C and detected with an RID‐10A refractive index detector. The 10‐HDA content was determined according to (Liu, Yang, Shi, & Peng, 2008) with a Shimadzu VP series liquid chromatograph, photo diode array detector, and an LC‐18 (5 μm) column (256 × 4.6 mm) with LC‐18 2CM precolumn KIT. Total protein content was determined using the Bradford (Bradford, 1976) method and BSA (bovine serum albumin) as the standard for calibration curve. Measurements at 595 nm were made on a Shimadzu UV Spectrophotometer.