Abstract For the first time, an attempt has been made to study the vitreous humor in situ by circular dichroism (CD). The vitreous, an avascular and acellular gel-like tissue, is optically transparent and homogeneous, and, thus, light scattering is minimal. The macromolecular components of this tissue, hyaluronate (HA), collagen and noncollagenous protein (NC-P), appear to exist in the matrix in a nonoriented fashion. As a result, no linear dichroism was observed. A typical CD of the vitreous shows a minimum at 206 nm with a shoulder at 220 nm and one small positive peak at ∼ 252 nm. Gaussian analysis resolves this spectrum into four component bands. CD analysis of individual components reveals that NC-P makes the major contribution to the dichroic strength of the vitreous; contributions of HA and collagen, on the other hand, are small. The positive peak arises largely from ascorbic acid in teh vitreous. CD measurement of the intact vitreous appears to be a useful technique for assessing the structure and changes of the constituent molecules in the normal and diseased vitreous.