Allium sativum L (garlic) is an essential component of many polyherbal oils used in traditional systems of medicine. Allyl disulfide has been a major component found in vegetable oil macerate of garlic, and can be used as reliable marker for determination of garlic in oil macerates of garlic. The HPLC separation of allyl disulfide was achieved on a Phenomenex Luna C18 (25 cm x 4.6 mm id x 5 pm particle size) column using acetonitrile-water-tetrahydrofuran (70 + 27 + 3, v/v/v) mobile phase at a flow rate of 1.0 mL/min. Quantitation was achieved with UV detection at 298 nm over the concentration range 8-48 microg/mL. HPTLC separation of allyl disulfide was achieved on an aluminum-backed layer of silica gel 60 F254 using n-hexane mobile phase. Quantitation was achieved by densitometric analysis at 298 nm over the 200-1200 ng/band concentration range. The methods were validated according to International Conference on Harmonization guidelines.