Objective <p>This study aimed to investigate the effectiveness of carotenoid-rich red palm olein (RPO) in treating or preventing xerophthalmia among Malaysian rural primary schoolchildren with confirmed vitamin A deficiency (VAD) (plasma retinol &lt;0.70 µmol/L) and marginal vitamin A status (plasma retinol 0.70 to &lt;1.05 µmol/L).</p> Methods <p>A double-blinded, cluster randomized controlled trial was conducted on 504 schoolchildren aged 8–12 years from ten rural primary schools in Malaysia. The selected schools were randomly assigned into two groups: the experimental group (<i>n</i> = 249) received RPO-enriched biscuits (326.3 µg retinol equivalents/day), while the control group (<i>n</i> = 255) received palm olein (PO)-enriched biscuits for 6 months (4 days/week). Chi-square test and mixed effects logistic regression model were applied to examine the effects of supplementation on the resolution or prevention of xerophthalmia.</p> Results <p>After 6-month supplementation, a higher resolution rate of xerophthalmia (31.2%) was observed in the experimental group compared to the control group (24.1%), but this difference lacked statistical significance (<i>p</i> = 0.11). Nevertheless, the experimental group showed a significantly higher prevention rate of xerophthalmia (81.8%) or conjunctival xerosis (82.5%) than the control group (56.5% or 57.6%, respectively) after 6-month supplementation (<i>p</i> &lt; 0.001). Moreover, schoolchildren in the experimental group were 4.8 times less likely to develop xerophthalmia compared to the control group.</p> Conclusion <p>A 6-month supplementation of RPO-enriched biscuits demonstrated potential benefits in preventing xerophthalmia or conjunctival xerosis among vitamin A-deficient primary schoolchildren in rural areas of Malaysia. Therefore, RPO may offer a promising preventive solution to xerophthalmia for populations at a high risk of developing VAD.</p> Clinical trial registration <p>ClinicalTrials.gov (NCT03256123).</p>