The purpose of this work was to develop an extraction technique to yield a betulinic acid-(BA) enriched extract of the traditional anti-anxiety plant Souroubea sympetala Gilg (Marcgraviaceae). Five extraction techniques were compared: supercritical carbon dioxide extraction (SCE), conventional solvent extraction with ethyl acetate (EtOAc), accelerated solvent extraction (ASE), ultrasonic assisted extraction (UAE) and soxhlet extraction (Sox). The EtOAc and SCE extraction methods resulted in BA-enriched extracts, with BA concentrations of 6.78 [1] 0.2 and 5.54 [1] 0.2 mg/g extract, respectively, as determined by HPLC-APCI-MS.The bioactivity of the BA-enriched extracts was compared in the elevated plus maze (EPM), a validated rodent anxiety behavior assay. Rats orally administered a 75 mg/kg dose of SCE extract exhibited anxiolysis as compared with vehicle controls, with a 50% increase in the percent time spent in the open arms, a 73% increase in unprotected head dips and a 42% decrease in percent time spent in the closed arms. No significant differences were observed between the SCE and EtOAc extracts for these measures, but the animals dosed with SCE extract had significantly more unprotected head dips than those dosed with the EtOAc extract. The SCE extract demonstrated a dose-response in the EPM, with a trend toward decreased anxiety at 25 mg/kg, and significant anxiolysis was only observed at 75 mg/kg dose. This study demonstrates that SCE can be used to generate a betulinic acid-enriched extract with significant anxiolysis in vivo. Further, the study provides a scientific basis for the ethnobotanical use of this traditional medicine and a promising lead for a natural health product to treat anxiety.
Martha Mullally,1† Kari Kramp,1,4† Chris Cayer,1,2 Ammar Saleem,1 Fida Ahmed,1 Calum McRae,3 John Baker,3 Andrew Goulah,4 Marco Otorola,5 Pablo Sanchez,5 Mario Garcia,5 Luis Poveda,5 Zul Merali,2 Tony Durst,1,6 Vance L. Trudeau7 and John Thor Arnason1*
1Centre for Research in Biopharmaceuticals and Biotechnology, Department of Biology, University of Ottawa, Ottawa, ON, Canada, K1N 6N5
2Institute of Mental Health Research, University of Ottawa, Ottawa, Ontario, Canada K1N 6N5
3Bioniche Life Sciences Inc., 231 Dundas Street East, Belleville, ON, Canada, K8N 1E2
4Loyalist College,Wallbridge-Loyalist Road, P.O. Box 4200, Belleville ON, Canada, K8N 5B9
5Universidad Nacional, Heredia, Costa Rica
6Department of Chemistry, University of Ottawa, Ottawa, ON, Canada, K1N 6N5
7Centre for Advanced Research in Environmental Genomics, Department of Biology, University of Ottawa, Ottawa, ON, Canada, K1N 6N5
PHYTOTHERAPY RESEARCH Phytother. Res. (2010) Published online inWiley InterScience (www.interscience.wiley.com) DOI: 10.1002/ptr.3246