The mineral status of sows plays a crucial role in their performance in the nursery and their productive career. Insufficient mineralisation is one of the factors implicated in lameness and leg issues, which degrade sow health and well-being, and reduce longevity in the herd. Mineral status can be monitored using urinary biomarkers assessed routinely in human medicine; however, free-catch sampling with a cup make it difficult to monitor large numbers of animals reared in groups. The aim of this study was to assess effects of using vaginal tampons to collect urine on the results of chemical composition analysis of mineral content and biomarkers of bone mineralisation. One spontaneous urination of each of 10 multiparous sows was sampled during the last week of gestation using four methods: control collection in a cup vs. collection with three models of commercially available tampons (i.e. A, B and C). Urine concentrations of phosphorus and calcium (minerals) and of two biomarkers of bone mineralisation (CTX-I and DPD) standardised by creatinine were determined. The sampling method did not influence the values of these minerals and biomarkers, except for calcium concentration (P < 0.001), which was lower using tampon B. This study suggests that it is possible to use vaginal tampons for urine sampling to analyse minerals and biomarkers of bone mineralisation in gestating sows. Tampon sampling is thus a way to routinely obtain urine from a large number of animals to ensure that they are monitored accurately.