Spd 20a ultraviolet detector

The molecular weights of polymers were determined by GPC using a liquid chromatography system equipped with a (RID-10A) differential refractive index detector (λ = 633 nm) and SPD-20A ultraviolet detector (Shimadzu). Samples were fractionated using 5.0 μm bead-size guard column (50 × 7.8 mm) and three Shodex KF-805L columns (300 × 8 mm, 10 μm n = bead-size, 5000 Å pore size) in series at 40 °C and eluted in N,N-dimethylacetamide (DMAC, HPLC grade, with 0.03% w/v LiBr) at a flow rate of 1 ml/min. A molecular weight calibration curve was produced using polystyrene standards ranging from 500 to 2 × 10⁶ Da.

The HMO mixture was prepared as described previously.^{7 (link)} Bacteria were grown in a medium containing HMO mixtures for 72 h. According to our previous report,^{7 (link)} the oligosaccharides remaining in the culture supernatant at the end of the experiment were labeled with p-aminobenzoic acid ethyl ester as an ultraviolet light-absorbing compound and quantified using HPLC. The labeled HMO components were separated using a Shimadzu Prominence HPLC system (Kyoto, Japan) with an L-column 2 ODS (Chemicals Evaluation and Research Institute, Tokyo, Japan). Modified HMOs were eluted with a 13:87 (vol:vol) mixture of acetonitrile and 100 mM ammonium acetate (pH 4.5) at 40°C and were detected using an SPD-20 A ultraviolet detector (Shimadzu) at 304 nm. The following oligosaccharides were used as controls: 2′-FL (Advanced Protein Technologies), 3-FL (Dextra Laboratories, cat L303), LNFP I (Dextra Laboratories, cat L502), LNDFH I (Dextra Laboratories, cat L602), LNDFH II (Dextra Laboratories, cat L603), LNT (IsoSep AB, cat 45/01–0010), lacto-N-neotetraose (IsoSep AB, cat 45/08–0010), DFL (IsoSep AB, cat 45/02–0010), LNFP II (IsoSep AB, cat 55/06–0001), LNFP III (IsoSep AB, cat 55/07–0005), and N-acetyl glucosamine (Nacalai Tesque, cat 00520–16).

To investigate changes in HDL particle size, patient HDL fractions were analyzed on an HPLC system equipped with a LC-20ADVP pump, a DGU-20A degassing unit, a CTO-20A column oven, an SPD-20A ultraviolet detector, and an SIL-20AC autoinjector (Shimadzu Corporation, Kyoto, Japan). Each sample (40 μL) was injected into serially connected size exclusion columns (PROTEIN KW-803 and KW-804; 300 mm × 8.0 mm i.d., Shodex, Tokyo, Japan), eluted with PBS at a flow rate of 1 mL/min, and monitored by absorbance at 280 nm.