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Introduction Prediabetes presents an opportunity for early intervention. Growing evidence suggests that psychological stress may contribute to glucose dysregulation, but the findings are inconsistent.This study aimed to clarify the association between perceived stress and glycemic measures, by first testing gender as a moderator, and then examining age as a moderator within each gender group.Research design and methods We analyzed cross-sectional data from 470 diabetes-free adults in Singapore. Participants completed the 10-item Perceived Stress Scale, comprising two subscales: perceived helplessness and perceived self-efficacy. Glycemic measures included fasting plasma glucose, glycated hemoglobin, and 2-hour plasma glucose (2h-PG) following an oral glucose tolerance test. Prediabetes was classified according to the American Diabetes Association diagnostic criteria.Results Multivariable regression analyses revealed significant moderating effects of gender on the relationship between perceived stress factors and both prediabetes status and 2h-PG levels. Specifically, higher perceived helplessness and perceived self-efficacy were significantly associated with a lower prevalence of prediabetes and lower 2h-PG levels among men. However, these associations were non-significant among women. Age significantly moderated the relationship between perceived helplessness (but not perceived self-efficacy) on prediabetes and 2h-PG levels in women; higher perceived helplessness was associated with a greater prevalence of prediabetes and higher 2h-PG levels among younger women.Conclusions Gender moderated the associations between perceived stress and both prediabetes prevalence and 2h-PG levels. Among women, age further moderated the association between perceived helplessness and these outcomes. Future research should consider both moderators. Tailored psychosocial stress management strategies may help reduce the prevalence of prediabetes and diabetes.Trial registration number NCT02838693.