Propionic acid

The samples were hydrolyzed in 0.2 M propionic acid (FUJIFILM Wako Pure Chemical Corporation) by incubating at 80 °C for 4 h. The hydrolysate was dried in a SpeedVac (Savant Instrument, Inc). For 1,2-diamino-4,5-methylenedioxybenzene-2HCl (DMB) derivatization, DMB solution was prepared as described previously (69 (link), 70 (link)). Twenty microliters of 0.01 M TFA and 20 μl of DMB solution was added to the dried samples, followed by incubating at 50 °C for 2 h. For the fluorometric HPLC analysis, the reaction mixture was diluted to 10 times and directly injected into a Handy ODS column (4.6 φ × 250 mm; FUJIFILM Wako Pure Chemical Corporation) and eluted with methanol/acetonitrile/0.05% TFA in water (4/6/90, v/v/v). A Jasco LC-900 HPLC system equipped with a Jasco FP-2025plus fluorescence detector (excitation, 373 nm; emission, 448 nm), operating at 1.0 ml/min at a column temperature of 26 °C, was used.

Powdered pullulan (PL) was purchased from Tokyo Chemical Industry Co., Ltd, Japan. Cotton lint cellulose was purchased from Walmart Stores, Inc. Trifluoroacetic anhydride (TFAA), acetic acid, propionic acid, and all other reagents were purchased from Wako Pure Chemical Industries, Ltd, Japan, and were used without further purification.

Propionic acid (PA), butyric acid (BA), L-glycine (Gly), L-alanine (Ala), L-serine (Ser), L-threonine (Thr), L-cysteine (Cys), L-methionine (Met), L-valine (Val), L-leucine (Leu), L-isoleucine (Ile), L-phenylalanine (Phe), L-tryptophan (Trp), L-aspartic acid (Asp), L-glutamic acid (Glu), L-asparagine (Asn), L-glutamine (Glu), L-histidine (His), L-asparagine monohydrate (Arg), L-tyrosine (Tyr), and L-ascorbic acid were purchased from FUJIFILM Wako (Osaka, Japan). Acetic acid (AA), L-lysine (Lys), and sodium butyrate were purchased from Nacalai Tesque (Kyoto, Japan), and proline (Pro) was purchased from Sigma-Aldrich (St. Louis, MO, USA). All chemicals were of analytical grade.