The composition of rhesus monkey aqueous humor has been studied in large-volume, pooled samples. Replicate determinations of the concentrations of a number of constituents have been carried out for both aqueous humor and serum from large veins by means of automatic analyzing equipment. Since aqueous humor has been obtained by anterior chamber paracentesis, it is a mixture of anterior and posterior chamber aqueous. When compared to serum, the pooled aqueous contains an excess of chloride, bicarbonate, ascorbate, lactate, uric acid, and several neutral amino acids. Rhesus monkey aqueous humor is deficient in calcium, urea nitrogen, phosphates, glucose, protein, creatinine, iron, bilirubin, cholesterol, triglycerides, a number of serum enzymes, acidic and basic amino acids, and several neutral amino acids. Sodium, potassium, magnesium, and two neutral amino acids (cysteine and valine) are of equal concentration in aqueous humor and serum. Glutathione concentration is very low in both aqueous humor and serum. Pooled rhesus monkey aqueous humor and serum are isosmolar, with measured osmolality being about 303 mOsm. Based upon the chemical analysis, a new solution has been formulated to substitute for primate aqueous humor during anterior ocular perfusion. This new solution causes very little change in the physiologic integrity of the outflow pathways during prolonged, repeated perfusion. In this respect, its effects are very similar to those of pooled rhesus monkey aqueous humor during perfusion of rhesus monkey eyes. In contrast, perfusion of rhesus monkey eyes with glutathione-bicarbonate-Ringer's solution has been shown to cause progressive increase of the total facility. To minimize physiologic alterations during operative procedures, a solution similar to this new one could be formulated for irrigation of the inside of the human eye.