Hyaluronic acid (HA) N-deacetylase(s) was quantified in whole. skin, using a novel method that involved reaction of skin with exogenous HA as substrate. Acetyl (CH3CO-) moieties generated were converted chemically to MeOAc and quantified using gas chromatography/mass spectrometry. HA (1.7 mg) and skin (1.0 g) yielded 3.32 and 769.00 mug of MeOAc from the 69.0- and 76.5-year-old-patient samples, respectively. Without added HA, 194.00 mug of product was obtained from the 76.5-year-old-patient samples. With chondroitin as substrate, the yields were 2.89 and 818.04 mug of MeOAc from the 69.0- and 76.5-year-old-patient samples, respectively. The K5 (capsular, Escherichia coli polysaccharide) substrate yielded no detectable product, except for 170.02 mug from the 76.5-year-old-patient samples. This highly sensitive method was used to demonstrate that human-skin-HA N-deacetylase(s) was first detectable at 69 years of age, highly active at 76.5 years of age, and specific for N-acetyl moieties Of D-GIcNAc and D-GalNAc where C-1 is beta-linked as in HA and CH.