Purpose This preliminary study examined the influence of menstrual cycle phase and hormone levels on acoustic measurements of vocal function in reproductive and postmenopausal females. Mean fundamental frequency (f0), speaking fundamental frequency (Sf0), and cepstral peak prominence (CPP) were evaluated. It was hypothesized that Sf0 and CPP would be lower during the luteal and ischemic phases of the menstrual cycle. Group differences with lower values in postmenopausal females and greater variability in the reproductive females were also hypothesized. Method A mixed factorial analysis of variance was used to examine differences between reproductive and postmenopausal females and the four phases of the menstrual cycle. Separate analyses of variances were implemented for each of the dependent measures. Twenty-eight female participants (15 reproductive cycling, 13 postmenopausal) completed the study. Participants were recorded reading the Rainbow Passage and sustaining the vowel /a/. Mean vocal f0, Sf0, and CPP were determined from the acoustic samples. Blood assays were used to determine estrogen, progesterone, testosterone, and neuropeptide Y levels at four data collection time points. Results Group differences in hormone levels and Sf0 values were established with the postmenopausal group having significantly lower hormone levels and significantly lower Sf0 than the reproductive cycling group across the phases. Analysis of the reproductive group by hormone levels and cycle phase revealed no significant differences for CPP or Sf0 across phases. Higher estrogen was identified in the ovulation phase, and higher progesterone was identified in the luteal phase. Conclusions Significant differences in hormone levels and Sf0 were identified between groups. Within the reproductive cycling group, the lack of significant difference in acoustic measures relative to hormone levels indicated that the measures taken may not have been sensitive enough to identify hormonally mediated vocal function changes. The participant selection may have biased the findings in that health conditions and medications that are known to influence voice function were used as exclusion criteria.