Background and Design: Excessive hair growth after isotretinoin treatment for acne vulgaris is not common, but may be one of the most undesirable side effects of the drug. The aim of this study is to investigate the relationship between systemic isotretinoin use and facial hypertrichosis. Materials and Methods: Female acne patients in premenopausal age were included in this prospective study. Laboratory tests [beta-human chorionic gonadotropin, total cholesterol, triglyceride, low-density lipoprotein, high-density lipoprotein, aspartate aminotransferase, alanine aminotransferase] were evaluated initially and monthly during the study period. Hormone levels [luteinizing hormone, follicle stimulating hormone, total testosterone, free testosterone, dehydroepiandrosterone-sulfate (DHEAS), prolactin, 17-hydroxyprogesterone, glucose, and insulin] and abdominopelvic/transvaginal ultrasonography were also evaluated when there was a complaint or clinical findings of excessive hair growth. Body mass index (BMI) was calculated at the beginning of the study. Severity of the acne was assessed with Global Evaluation Acne Scale (GEAS). Hirsutism scores were calculated with Modified Ferriman-Gallwey score (m-FGS). Baseline and monthly taken digital dermoscopic photographs from the chin and cheeks were transferred to the ImageJ program to count the hair. Hair increases of >5% at the end of the treatment according to the basal hair count was accepted as hypertrichosis. Results: Thirty patients aged between 18-34 (median: 21.5) participated in the study. Mean duration of the therapy was 6.2±0.6 months. Facial hair growth was detected in three (10%) patients. One patient had an elevated DHEAS level with normal abdominal ultrasonography findings. Without the cessation of isotretinoin therapy, DHEAS level decreased to normal limits after two months. There was no statistically significant difference found between the mean GEAS (p=0.52), basal m-FGS (p=0.42), and BMI (p=0.71) of three patients with facial hypertrichosis, and in the remaining 27 patients. Facial hypertrichosis disappeared spontaneously 2 months (1-3 month) after the treatment courses were completed. Conclusion: The patho-mechanism of isotretinoin induced facial hair growth is not fully clarified. Since the facial hypertrichosis disappeared spontaneously when the treatment was ended, we think that this may be due to a temporary drug induced hormonal imbalance in susceptible individuals.