Sodium borodeuteride

The total cell N-glycome and O-glycome of cell lines were prepared as described previously (60 , 61 (link)). O-Glycans were reduced with sodium borohydride or sodium borodeuteride (Sigma). Released glycans were permethylated with iodomethane and powdered sodium hydroxide (36 (link), 62 ). Permethylated glycans were fractionated by Sep-Pak C18 Classic Cartridge into 15, 35, 50, and 75% acetonitrile fractions and analyzed by MALDI-TOF and MALDI-TOF-TOF on an ABSciex 5800, as described previously (36 (link)). Glycan compositional assignment was confirmed by extensive MS/MS fragmentation analysis with manual interpretation and reconstruction of the precursor ions using GlycoWorkBench (63 (link)).

Hull-less barley was cultivated in Qinghai Province, China. Enzymes (thermostable α-amylase, amyloglucosidase and papin) were provided by Aladdin Biochemical Technology Co., Ltd. (Shanghai, China). The β-glucan assay kit was from Megazyme International (Wicklow, Ireland). d-xylose was obtained from J&K Scientific Ltd. (Beijing, China). Other monosaccharide standards (d-arabinose, d-galactose, d-glucose, glucuronic acid, and galacturonic acid), deuterium oxide (D₂O), dimethyl sulfoxide-d₆ (DMSO-d₆), methyl iodide, and sodium borodeuteride (98 atom% d) were bought from Sigma-Aldrich Chemical Co. (St. Louis, MO, USA). All other used reagents were of analytical grade unless otherwise specified.

Intestinal permeability was determined by quantification of urinary excreted sugars after ingestion of an oral sugar cocktail [42 (link)–44 (link)]. The oral sugar cocktail was taken by the HC subjects seven days after the endoscopic sigmoid colon mucosa and fecal specimens were collected. Fasted HC subjects were given an oral cocktail of 40 g sucrose, 2 g mannitol, 7.5 g lactulose mixed in 8 oz. of water and 1 g sucralose in two capsules at 6 AM, and then urine was collected for 24 hours. Urinary concentrations of excreted sugars were determined using gas chromatography and an internal standard as previously described [42 (link),44 (link)]. The intestinal permeability results were expressed as percent excretion of oral dose of the various sugars as previously described [42 (link),44 (link)].
Lactulose (4-o-β-D-galactopyranosyl-D-fructofuranose) was obtained from Bertek Pharmaceuticals, as brand name Kristalose. Mannitol (D-mannitol) and sucrose (α-D-glucopyranosyl-β-D-fructofuranoside) were obtained from Sigma Aldrich. Sucralose (1,6-dichloro-1,6-dideoxy-β-D-fructofuranosyl-4-chloro-4-deoxy-α-D-glucopyranoside) was supplied by Tate and Lyle. Trifluoroacetic acid ammonia, sodium borodeuteride, acetone, acetic anhydride and glacial acetic acid were purchased at their highest grade purity from Sigma Aldrich.

Dendrobium nobile Lindl. was obtained from Guizhou Chishui Guoli Dendrobium nobile Lindl. Development Co., Ltd. (Guizhou, China), dried and crushed, and then screened with 80 meshes.
MD2-IN-1, TLR4-IN-C34, IL-10, IL-6, IL-1 β, TNF-α ELISA kit were purchased from Shanghai Jianglai Biotechnology Co., Ltd. (Shanghai, China); mouse macrophage RAW264.7 was provided by Zhejiang Meisen Cell Technology Co., Ltd. (Wenzhou, China); DMEM High-Glucose Medium and FBS were acquired from Gibco (New York, NY, USA); CCK-8 Kit was purchased from Wuhan Doctor De Bioengineering Co., Ltd. (Wuhan, China); NO detection kit was obtained from Shanghai Biyuntian Biotechnology Co., Ltd. (Shanghai, China); trifluoroacetic acid, acetic acid, LPS, dimethyl sulfoxide, sodium borodeuteride, and acetic anhydride were purchased from Sigma (Kalamazoo, MI, USA).