This study aimed to compare three methods of predicting ovulation day: (1) a positive urinary luteinising hormone test (LH), (2) a sustained rise in salivary progesterone above critical difference (SP), and (3) a countback regression equation (CB), to determine variability in the menstrual cycle (MC) lengths and reproductive hormone concentrations of professional female soccer players. Eight players provided daily morning saliva samples for three consecutive cycles. Samples were analysed for oestradiol and progesterone concentrations. Each MC was separated into the follicular (FP) and luteal (LP) phases relative to the day of ovulation, using the three different methods. MC length ranged from 24 to 32 days (28.3 ± 2.4 days); intra-assay coefficient of variation (7.5%) exceeded inter-assay coefficient of variation (4.6%). Ovulation estimated using SP (15.4 ± 3.0 days) occurred later than LH (13.3 ± 2.0 days) (P = 0.017). The CB method (14.1 ± 1.8 days) did not differ from SP (P = 0.102) or LH (P = 0.262). Oestradiol and progesterone levels varied significantly between sub-phases (P < 0.001). Inter-variability surpassed intra-variability for both hormones. Differences in methods for predicting ovulation indicate the need for standardised protocols. Individual variation in MC length and hormone concentrations challenges the narrative for group-level MC recommendations, emphasising the need for individualised hormone monitoring across multiple cycles.
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