Background Certain ovarian cancers previously more common in postmenopausal women are now increasingly observed in women of reproductive age. The research on using β-HCG as a diagnostic biomarker for ovarian cancer in women of reproductive age is ongoing. This particular study assessed the level of serum β-HCG in non-pregnant women of reproductive age and determined its potential association with suspicious ovarian ultrasonography results in Port Harcourt, Nigeria. Material and methods This study utilized a descriptive-analytic design on a quota sample of 224 diagnostic case notes of women aged 18-40. Data collection involved a data extraction form. Data analysis employed descriptive statistics, Chi-square, Fisher's exact test, and Odds Ratio at 95% confidence and 5% significance levels. Results About 5.8% of the participants exhibited detectable levels of serum β-HCG above 5 IU/L (World Health Organization reference) at a mean concentration of 5.87 (±1.75) IU/L. About 4% of the participants had suspicious ovarian lesions identified through ultrasonography. Participants with elevated serum β-HCG levels above the WHO reference were 59 times more likely to have suspicious ovarian lesions, with an odds ratio of 59.4 (95%CI: 12.3–287.8, p = 0.001). There was a significant association between serum β-HCG level and age (p = 0.041) as well as parity (p < 0.001). Conclusions This study demonstrated that Serum β-HCG levels above the WHO reference were associated with suspicious ovarian lesions. Non-pregnant women should undergo serum β-HCG testing at least yearly to facilitate the early detection of ovarian anomalies.