Journal
JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES
Volume 857, Issue 2, Pages 210-218Publisher
ELSEVIER
DOI: 10.1016/j.jchromb.2007.07.017
Keywords
size-exclusion HPLC; reversed-phase HPLC; protein purification; qualitative analysis; quantitative analysis; method validation; fractionation; enamel matrix derivative; emdogain
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High performance liquid chromatography (HPLC) methods were used to analyse a 5 kDa component purified from enamel matrix derivative (EMD), the active ingredient in Emdogain((R)), a commercial product for periodontal tissue regeneration. After initial purification by size-exclusion chromatography (SEC) on a 100 cm x 5 cm column (Bio-Gel P-30 Fine, 280 nm), collected fractions were analysed by size-exclusion HPLC (SE HPLC; TSK-Gel Super SW2000, 220 nm). The fractions containing only the 5 kDa component were analysed by reversed-phase high-pressure chromatography (RP HPLC; YMC-Pack ODS-A, 200 nm), revealing four peaks of the 5 kDa component. From 1200 mg of EMD (of which 9% is the 5 kDa component), approximately 65 mg of lyophilised 5 kDa component were obtained, corresponding to a recovery of 60%. The SE HPLC method was mainly suitable for qualitative analysis, whereas the RP HPLC method was appropriate for both qualitative and quantitative analysis. (c) 2007 Elsevier B.V. All rights reserved.
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