MSACL 2025 Abstract

Assessment of Serum 3-Epi-25-Hydroxyvitamin D3, Total 25-Hydroxyvitamin D3, and 25-Hydroxyvitamin D2 in the Korean Population with UPLC-MS/MS

Sung-Eun Cho, M.D.(1), Jungsun Han, M.S.(1), Rihwa Choi, M.D.(1), Sang Gon Lee, M.D.(1), Hyung-Doo Park, M.D.(2)
(1) GC Labs, Gyeonggi-do, Korea, (2) Department of Laboratory Medicine and Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea

Sung-Eun Cho, M.D., Ph.D. (Presenter) GCLabs ESAC

Relevant Financial Disclosures (within past 24 months, reported on Apr 20, 2026) No relevant financial relationship(s) to disclose. Conflict mitigation NOT REQUIRED.

INTRODUCTION:
The liquid chromatography-tandem mass spectrometry method (LC-MS/MS) identified an epimeric form of 25-hydroxyvitamin D3 (3-epi-25-OH-D3) concentrations. The determination of 3-epi-25-OH-D3 has been nationally surveyed in the USA’s National Health and Nutrition Examination Survey (NHANES) since 2007 using LC-MS/MS method. In Korea, the 3-epi-25-OH-D3 was included for the first time in the Korea National Health and Nutrition Examination Survey (KNHANES) IX (from 2022 to 2024). We conducted this study and measured the 3-epi-25-OH-D3 concentrations, and now we evaluated the results of 3-epi-25-OH-D3 in KNHANES IX to find out its clinical status in Korea.

METHODS:
The UPLC-MS/MS was used for the measurement of 3-epi-25-OH-D3, 25-hydroxyvitamin D3 (25-OH-D3), and 25-hydroxyvitamin D2 (25-OH-D2). We used Nexera-X2-LC-30AD UPLC (Shimadzu, Japan) system equipped with a Kinetex XB C18 column (2.1X150 mm, 2.6 um; Phenomenex, USA) on a Triple Quad 4500MD (Sciex, USA) MS/MS system. This method had a longer run time of 16 minutes to separate 3-epi-25-OH-D3. We evaluated the above vitamin Ds based on gender and age group and then compared them with other markers.

RESULTS:
The concentration ranges of 3-epi-25-OH-D3, 25-OH-D3 and 25-OH-D2 in 11,744 Korean participants were 0.50 (lower limit of quantification (LLOQ))-15.66, 2.32-116.78, 0.30 (LLOQ)-22.13 ng/mL, and mean±SD concentrations were 1.15±0.85, 22.60±10.60, and 0.44±0.62 ng/mL, respectively. The concentration of 3-epi-25-OH-D3 was not significantly different between males and females. The blood concentrations of 25-OH-D metabolites differed across age groups in both males and females (P < 0.0001). The concentrations of 25-OH-D2, 3-epi-25-OH-D3, 25-OH-D3, and total 25-OH-D (which is the sum of 25-OH-D2, 3-epi-25-OH-D3, and 25-OH-D3) were highest in the 60s, followed by higher concentrations in the age group of 70s and above. In those aged group under 60, the concentrations tended to increase according to the age increase. There was a highly significant correlation between 3-epi-25-OH-D3 and total 25-OH-D3 (sum of 3-epi-25-OH-D3 and 25-OH-D3) (Pearsons’s correlation coefficient r=0.74, R2=0.5404, slope=0.0587, P<0.0001). The minimum and maximum % 3-epi-25-OH-D3 were 1.35 and 31.47 %, respectively. Although the mean 3-epi-25-OH-D3 concentration increased linearly by quartile of serum total 25-OH-D3 concentration, the % 3-epi-25-OH-D3 was highest in the Q1 quartile of total 25-OH-D3 concentration (P<0.001). The inclusion of 3-epi-25-OH-D3 concentration in the estimate of serum total 25-OH-D reduced the prevalence of serum 25-OH-D deficiency, insufficiency, and suboptimal level by 1.3, 3.2, and 3.8 percentage points, respectively. The prevalence of serum total 25-OH-D > 70 ng/mL increased from 0.2 to 0.3 % when the 3-epi-25-OH-D3 concentration was included in the total estimate. Every marker except hsCRP had significant correlations with % 3-epi-25-OH-D3. Among them, PLT, BUN, Total cholesterol, HDL cholesterol, and LDL cholesterol showed a negative correlation with % 3-epi-25-OH-D3.

CONCLUSION:
This is the first large-scale study of 3-epi-25-OH-D3 serum concentrations across all age groups in the Korean population. Accurate measurement of 3-epi-25-OH-D3 concentration is necessary for the reliable results of vitamin D metabolites in KNHANES of Korea.

KEYWORDS: Tandem mass spectrometry, 3-epi-25-OH-D3, KNHANES