The surface lipids of Nicotiana benthamiana contained novel glycerolipids and several varieties of glycolipids. As glycerolipids, the triacylglycerol, 1,3-diacylglycerol, and 1,2-diacylglycerol types of glycerolipids were isolated and identified. Each lipid contained acetyl, 16-methylheptadecanoyl, and 18-methylnonadecanoyl moieties. The acetylated position of each lipid was determined by 2D-NMR, using the HMBC technique. The structures were 1,3-di-O-acetyl-2-0-acylglycerol, 1-0-acetyl-3-0-acylglycerol, and 1-0-acetyl-2-0-acylglycerol. As glycolipids, one glucose ester and four types of sucrose esters were isolated and identified. These glycolipids contained acetic acid and such branched short-chain fatty acids as 5-methylhexanoic, 4-methylhexanoic, 6-methylheptanoic, and 5-methylheptanoic acids. The structure of the glucose ester was 3,4-di-O-acyl-iX-o-glucopyranose. The structures of the sucrose esters were 6-0-acetyl-4-0-acyl-iX-o-glucopyranosyl-(3-0-acyl)-fJ-o-fructofuranoside, 4-0-acyl-iX-o-glucopyranosyl-(3- O-acyl)-fJ-o-fructofuranoside, 3,4-di-O-acyl-iX-o-glucopyranosyl-fJ-o-fructofuranoside, and 6-0-acetyl-iXo-glucopyranosyl-fJ-o-fructofuranoside.