Journal
JOURNAL OF SEPARATION SCIENCE
Volume 35, Issue 23, Pages 3281-3292Publisher
WILEY-V C H VERLAG GMBH
DOI: 10.1002/jssc.201200531
Keywords
Gel filtration; High-speed counter-current chromatography; Preparative high-performance liquid chromatography; Soyasaponins
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Funding
- National Natural Science Foundation of China [31071630/C200303]
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Soyasaponins are naturally occurring triterpenoid glycosides associated with many biological activities. The aim of the present study was to develop an effective method for isolation and purification of differently glycosylated, acetylated, and 2,3-dihydro-2,5-dihydroxy-6-methyl-4H-pyran-4-one (DDMP)-conjugated soyasaponins from soy hypocotyls. Both gel filtration using Sephadex LH-20 chromatography (Amersham Pharmacia Biotech AB; elution phase: methanol, flow rate: 3.0 mL/min, sample loading: 60 mg) and high-speed countercurrent chromatography (stationary phase: n-butanolacetic acid (5.0%, v/v), mobile phase: water flow rate: 3.0 mL/min, sample loading: 100 mg) could effectively fractionate isoflavones and soyasaponins from the crude extract with yield of soyasaponin complexes 20.5 mg and 22.3 mg, respectively. After fractionation, the soyasaponin complexes could be purified further using preparative HPLC to separate individuals. A total of nine soyasaponins, triacetyl soyasaponin Ab (yield 1.55%, HPLC purity >98%), Aa (2.68%, >99%), Ab (18.53%, >98%), Ae (0.85%, >98%), Ba (0.63%, >91%), Af (1.12%, >85%), Bb (3.45%, >98%) and Be (0.59%, >76.8%) were obtained. DDMP-conjugated groups, ag (2.06%, >85%), beta g (7.59%, >85%), and ?g (0.29%, >85%) that were very labile even in mild conditions, were also collected. The method described here can be used as an effective protocol to separate different soyasaponins occurring in the original sample.
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