Sil 10a

1 H and 13 C NMR spectra were acquired in CDCl3 on a Varian 500 NMR spectrometer at room temperature. The molecular weights of the products were measured by gel permeation chromatography (GPC) in chloroform on a Shimadzu SEC system (CBM-20A, SPD-10AVP, SIL-10A, LC-10ATVP, FCV-10ALVP, CTO-10AVP, RID-10A, and FRC-10A, Shimadzu, Japan).
Sample solutions were passed through a syringe filter (Sartorius Stedim, Minisart RC 4 or RC 15; pore size 0.45 µm) before GPC analysis. Shodex columns (K802, K802.5, and K805) with a guard column (Shodex, K-G) were used. Number-and weight-averaged molecular weights (Mn and Mw) and polydispersity indices (Mw/Mn) were estimated using polystyrene standards (Shodex).
Matrix-assisted laser desorption/ionization time-of-flight mass spectra (MALDI-TOF MS) were recorded on a Bruker Autoflex III machine in the positive ion linear mode. 2,5-Dihydroxybenzoic acid (DHB) was used as a matrix for these measurements.

Organic acids were determined by HPLC according to Laye et al. (1993) and Fernandez-Garcia and Mc-Gregor (1994) with some modifications. Five milliliters of homogenized kefir samples was added to 25 mL of 0.01 N H 2 SO 4 , vortexed for 1 min, and then centrifuged (Sigma 3K30, Germany) at 7,000 × g at 4°C for 7 min. The resulting supernatants were passed through a syringe filter (pore size = 45 μm; Chromafil GF/ PET-45/25, Macherey-Nagel AG, Duren, Germany). Filtered samples were injected into a Shimadzu HPLC system equipped with a pump (Shimadzu LC-20AT), photodiode array detector (Shimadzu SPD-M20A), column oven (Shimadzu CTO-10AS VP) set at 65°C, auto sampler (Shimadzu SIL-10A), and data station (Shimadzu CBM-20A). The analyses were carried out at a flow rate 0.7 mL/min with a Shim-Pack column (150 × 4.6 mm, 4 μm; Tokyo, Japan) using 10 mM H 2 SO 4 as the mobile phase. Organic acids were defined using external standards (lactic, acetic, citric and butyric acids; Sigma-Aldrich). The quantification of the results was performed depending on standard curves of individual organic acids covering the concentration range of 10 to 500 mL/L.

Molecular weights of HePS-1HePS-6 were determined by size-exclusion chromatography (SEC) as described [36] , using a solvent delivery system (LC-10AD), autosampler (SIL-10A), RI detector (RID-10A; all Shimadzu, Kyoto, Japan) and a size-exclusion column (Shodex OH-PK SB-805 HQ 300 × 8 mm, Showa Denko, Tokyo, Japan). Dextran standards of M w 4.5 MDa, 1.45 MDa, 560 kDa, 350 kDa (American Polymer Standards Corporation, Mentor, OH, USA), 276.5 kDa, 196.3 kDa, 123 kDa, 43 kDa (Pharmacosmos, Holbaek, Denmark), and pullulan of 22 kDa were used for calibration. Standards (1-2.7 mg/ml) and HePSs (1-2 mg/ml) were dissolved in mobile phase (10 mM sodium citrate/citric acid pH 4.0), degassed, kept overnight, filtered (0.45 µm filters; Frisenette ApS, Knebel, Denmark), and 100 µl of the solution was subjected to SEC at a flow rate of 0.5 ml/min. The SEC data were analyzed by TriSEC conventional GPC software.