Osteoporosis is highly prevalent, particularly in developing countries. However, bone turnover marker reference ranges for management of osteoporosis in Asian population are yet to be explored and established. Thus, this study aims to develop a regional bone turnover markers (BTMs) reference database by combining country-specific reference database from five ASEAN countries: Malaysia, the Philippines, Singapore, Thailand, and Vietnam. We established a healthy reference population of 746 healthy premenopausal women aged 20 to 44 years old. Serum Procollagen 1 N-Terminal Propeptide (P1NP), Osteocalcin (OC), and Beta-Crosslaps (CTX) concentrations were measured using an automated immunoassay analyzer system, the cobas® modular analyzer systems (Roche Diagnostic Gmbh). The reference interval was defined as the central 95 % range. The estimated reference interval for CTX was 128 to 811 ng/L, OC was 9.0 to 33.0 μg/L, and for P1NP, the range was 22.8 to 96.5 μg/L. Comparison across countries showed that Singaporeans had the highest levels of median CTX along with Thais and Filipinos, who had significantly higher levels of P1NP and OC. Exploratory analysis on the associations with age showed that BTMs decreased with increasing age at 20 to 29 years old and plateaued after 30 years old. When excluding participants in their 20s, the reference interval estimated were CTX: 117-678 ng/L, P1NP: 21.6-85.8 μg/L and OC: 3.5-27.0 μg/L respectively. To the best of our knowledge, this is the first study to report BTMs reference intervals based on a healthy premenopausal Southeast Asian population which will contribute to the appropriate assessment and monitoring of bone turnover rate in the evaluation and management of osteoporosis in the Southeast Asian region. LAY SUMMARY: Osteoporosis is a common health issue, especially in developing countries. However, there is a lack of information on bone health markers specific to the Southeast Asian population. This study aimed to fill this gap by creating a reference database for bone turnover markers (BTMs) in Southeast Asian countries, including Malaysia, the Philippines, Singapore, Thailand, and Vietnam. The researchers studied 746 healthy women aged 20 to 44 years and measured blood markers related to bone health. The reference interval, representing the normal range, was determined. For example, the normal range for CTX was found to be 128 to 811 ng/L, for Osteocalcin was 9.0 to 33.0 μg/L, and for P1NP, the range was 22.8 to 96.5 μg/L. When excluding participants in their 20s, the reference intervals estimated were CTX: 117-678 ng/L, P1NP: 21.6-85.8 μg/L and OC: 3.5-27.0 μg/L respectively. Comparing the results across countries, Singaporeans, Thais, and Filipinos showed variations in their biochemical bone marker levels. Additionally, the study observed changes in the levels with age, with a decrease in BTMs observed after the age of 30. This groundbreaking study provides the first-ever reference intervals for BTMs in a healthy premenopausal Southeast Asian population. These findings will help in the proper assessment and monitoring of bone health, contributing to the management of osteoporosis in the Southeast Asian region.