We demonstrate a method for estimating absorption and backscattering coefficients by inverting glider-measured profiles of the downwelling irradiance and upwelling radiance. The inversion method was validated against approximately 1,300 profiles of data from 22 glider missions within the Gulf of Maine over a 10 year period. The backscattering coefficient at 532 nm was estimated with a mean absolute error of 21% and bias of 0.01% compared to measured values. We could only quantitatively evaluate the absorption coefficient against the fluorometry data, but found that profiles of fluorescence and absorption were in quantitative agreement. With absorption and backscattering coefficients acting as a basis for studying the biogeochemical parameters of the constituents in the water column, these results show the potential of bio-optical gliders for studying marine ecosystems under varying sky conditions.