Bsa lyophilized powder

The bioactive Lf from bovine whey was kindly supplied by the Tatua Cooperative Dairy Company Ltd (New Zealand). Poly-L-Lysine hydrochloride (PLL, Mw 15,000–30,000), BSA lyophilized powder (≥96%), TA (ACS grade), BSA conjugated with fluorescein isothiocyanate (BSA/FITC), bile salts (for microbiology), pancreatin from porcine pancreas (≥100 USP U/mg), pepsin from porcine gastric mucosa (3802 U/mg), mini protease inhibitor cocktail cOmplete™, hydrochloric acid, calcium chloride dehydrate, sodium bicarbonate, sodium chloride, sodium hydroxide, acetonitrile (for HPLC, ≥99.9%) were purchased from Sigma-Aldrich. Anhydrous sodium carbonate was purchased from Alfa Aesar. 0.1% aqueous solution of trifluoroacetic acid (TFA, LC/MS grade, ≥99.99%) was purchased from VWR Chemicals. DQ™ Red BSA was purchased from Molecular Probes Inc, USA. The bovine lactoferrin ELISA kit was purchased from Bethyl Laboratories, Inc., USA. Cyanine 7 N-hydroxysuccinimide ester (Cy7-NHS) was purchased from Lumiprobe Corporation, USA. Cy7-labelled Lf (Cy7-Lf) has been prepared according to a standard protocol provided by Lumiprobe Co and purified by dialysis. All chemicals were used as received without further purification. Deionized (DI) water (specific resistivity higher than 18.2 MΩcm) from Milli-Q plus 185 (Millipore) water purification system was used to make all solutions.

ALG, BSA lyophilized powder, sodium bicarbonate, 2,4,6-trinitrobenzenesulfonic acid (TNBS), CaCl₂ and 4-(2-Hydroxyethyl) piperazine-1-ethanesulfonic acid sodium salt (HEPES), CIP and IND were all purchased from Sigma-Aldrich, Dorset, UK. Sodium dodecyl sulphate was purchased from ACROS, Leicestershire, UK.
The British Pharmacopeia standard was used to prepare the simulated gastric medium (SGM) and simulated intestinal medium (SIM) as follows:
-

SGM: 0.1M hydrochloric acid (pH 1.2).

-

SIM: 2.4g HEPES and 8.5g NaCl were dissolved in 1 L distilled water to obtain concentrations of 9.2 mM and 150 mM respectively. The pH was then adjusted to 6.5 using diluted HCl or NaOH solutions as required.