Background: Genitourinary syndrome of menopause (GSM) is a chronic and progressive syndrome that markedly impairs sexual quality of life in postmenopausal women. While vaginal laser therapy has emerged internationally as a promising non-hormonal treatment, data from Asian populations remain limited. Aim: To evaluate the effects of fractional CO2 vaginal laser therapy on sexual quality of life in Taiwanese women with GSM, with subgroup analysis by recent hormone replacement therapy (HRT) exposure. Methods: This retrospective single-center study included 27 postmenopausal women treated at a secondary teaching hospital in Taiwan from August 2022 to September 2024. All underwent three sessions of SmartXide² CO2 vaginal laser therapy. Sexual quality of life was assessed with the Pelvic Organ Prolapse/ Urinary Incontinence Sexual Questionnaire short form (PISQ-12, item 5-8) and a Sexual Satisfaction Visual Analog Scale (VAS) at baseline, 3, 6, and 12 months. Outcomes: The primary outcome was improvement in PISQ-12 short form scores and Sexual Satisfaction VAS score across the study period, with subgroup analysis by HRT status. Results: The mean PISQ-12 short form score improved from 9.9 ± 4.7 to 12.7 ± 3.5 (p = 0.004). Significant improvement was observed in the non-HRT group (p = 0.008), but not in the HRT group (p = 0.154). In longitudinal analysis, the non-HRT group demonstrated sustained increases in Sexual Satisfaction VAS score up to 12 months (all p < 0.01 vs baseline), whereas the HRT group showed only transient, non-significant changes. Clinical Implications: Fractional CO2 vaginal laser therapy may serve as a valuable non-hormonal option for GSM, particularly for women who are not candidates for or do not respond well to HRT. Strengths & Limitations: This study used a standardized laser protocol in an Asian cohort, enhancing validity and local relevance, but its retrospective single-center design, small sample size, self-reported measures, and short follow-up limit generalizability. Conclusions: Fractional CO2 vaginal laser therapy improved sexual quality of life in postmenopausal women with GSM, with sustained benefits most evident in those without recent HRT exposure. It may represent a valuable alternative for patients who are not candidates for or do not respond to hormone therapy.