Measuring Vitamin K1 Concentrations in Dogs with Chronic Enteropathy Using LC–MS/MS

As information regarding measurement and supplementation of vitamin K1 (vitK1) in dogs with chronic enteropathy (CE) is currently limited, a joint study conducted by the University of Tennessee (Knoxville, Tennessee) and the University of Pennsylvania School of Veterinary Medicine (Philadelphia, Pennsylvania) set out to compare vitK1 concentrations of healthy dogs to dogs with CE and determine if supplementation with vitK1 increases vitK1 concentrations compared to a placebo. Vitamin K concentrations were measured before and after supplementation using liquid chromatography tandem mass spectrometry (LC–MS/MS) and compared to vitK1 concentrations in the healthy cohort. A paper based on this study was published in the Journal of Veterinary Internal Medicine (1).

CE in dogs encompasses a variety of persistent, debilitating intestinal diseases that may result in intestinal barrier dysfunction, altered intestinal immunity, and nutrient malabsorption; for a diagnosis of CE to be made, gastrointestinal (GI) signs (for example, vomiting, diarrhea, regurgitation, flatulence) must be present for over three weeks, and other potential causes (such as parasites, neoplasia, endocrinopathy) must be ruled out (2–4). The initial diagnosis can be refined through diet trials or immunosuppressive treatments (food-responsive and immunosuppressant-responsive enteropathies, respectively) (5). If CE is associated with protein-losing enteropathy (PLE), the loss of proteins, lymphocytes, and lipids can be clinically relevant and potentially life-threatening (6).

Accurately assessing vitamin K1 status is challenging, as direct measurement is analytically difficult and often impractical due to its rapid utilization and minimal storage in the body (7). While vitK1 can be measured in humans, it is markedly affected by diet, meal pattern, and the presence of triglycerides; it also requires specialized laboratories and equipment for optimization because of extremely low circulating concentrations (8). As a result of these limitations, the measurement of vitK1 functional activity is now considered to be a more accurate reflection of vitK1 status in humans (9,10). While functional activity can be assessed in humans by determining the percentage of undercarboxylated osteocalcin (%ucOC), which is a sensitive functional marker of vitK1 (11), there are no currently available validated tests for %ucOC in dogs; direct measurement is achieved using LC–MS/MS (12).

The researchers report that they did not find a difference in serum vitamin K concentrations between healthy dogs and dogs with CE (before supplementation) using LC–MS/MS. The most plausible reason for this is that the lowest concentrations quantifiable by LC–MS/MS are above the actual difference between the groups, thereby showing that LC–MS/MS may not be the most sensitive method for vitK1 detection (1).

The researchers acknowledge several limitations in their study, including the small number of dogs enrolled and the lack of standardization of treatments for CE, which causes difficulty in interpreting the benefits of vitK1 treatment alone. Improvements in some of the clinical markers evaluated (such as albumin, canine inflammatory bowel disease activity index [CIBDAI] scores, and cholesterol) could be due to a combination of vitK1 supplementation and the animal's other prescribed treatments. Another limitation reported was measuring healthy control dog vitK1 concentration at only one time point. Normal variations may occur in vitK concentrations, and including additional measurements would have been useful to investigate such deviations. Also, the lowest limit of detection of vitK1 using LC–MS/MS was 0.1 ng/mL, which was the result obtained for dogs with chronic enteropathy supplemented with a placebo as well as healthy dogs. This method of measuring vitK1, therefore, may not be sensitive enough to detect differences. Based on these limitations, the researchers believe that future studies should use standardized treatment protocols in all groups in conjunction with vitK1 supplementation (1).

Group of twelve dogs. © Erik Lam - stock.adobe.com

References

1. Smith, J. M.; Smith, C. K.; Zhu, X.; et al. Supplementation of Vitamin K1 in Dogs with Chronic Enteropathy. J. Vet. Intern Med. 2025, 39 (3), e70111. DOI: 10.1111/jvim.70111

2. Dandrieux, J. R. Inflammatory Bowel Disease Versus Chronic Enteropathy in Dogs: Are They One and the Same? J. Small Anim. Pract. 2016, 57 (11), 589–599. DOI: 10.1111/jsap.12588

3. Washabau, R. J.; Day, M. J. Canine and Feline Gastroenterology. Elsevier Health Sciences, 2012.

4. Dandrieux, J. R. S.; Mansfield, C. S. Chronic Enteropathy in Canines: Prevalence, Impact and Management Strategies. Vet. Med. (Auckl) 2019, 10, 203–214. DOI: 10.2147/VMRR.S162774

5. Jergens, A. E.; Heilmann, R. M. Canine Chronic Enteropathy-Current State-of-the-Art and Emerging Concepts. Front Vet Sci. 2022, 9, 923013. DOI: 10.3389/fvets.2022.923013

6. Craven, M. D.; Washabau, R. J. Comparative Pathophysiology and Management of Protein-Losing Enteropathy. J. Vet. Intern. Med. 2019, 33 (2), 383–402. DOI: 10.1111/jvim.15406

7. Shearer, M. J.; Newman, P. Recent Trends in the Metabolism and Cell Biology of Vitamin K with Special Reference to Vitamin K Cycling and MK-4 Biosynthesis. J. Lipid Res. 2014, 55 (3), 345–362. DOI: 10.1194/jlr.R045559

8. Fusaro, M.; Gallieni, M.; Rizzo, M. A.; et al. (2017). Vitamin K Plasma Levels Determination in Human Health. Clin. Chem. Lab. Med. 2017, 55 (6), 789–799. DOI: 10.1515/cclm-2016-0783

9. Shea, M. K.; Booth S. L. Concepts and Controversies in Evaluating Vitamin K Status in Population-Based Studies. Nutrients 2016, 8 (1), 8. DOI: 10.3390/nu8010008

10. Viggiano, D.; Wagner, C. A.; Blankestijn, P. J.; et al. Mild Cognitive Impairment and Kidney Disease: Clinical Aspects. Nephrol. Dial. Transplant. 2020, 35 (1), 10–17. DOI: 10.1093/ndt/gfz051

11. McKeown, N. M.; Jacques, P. F.; Gundberg, C. M.; et al. Dietary and Nondietary Determinants of Vitamin K Biochemical Measures in Nen and Women. J. Nutr. 2002, 132 (6), 1329–1334. DOI: 10.1093/jn/132.6.1329

12. Ma, Y.; Yu, S.; Mu, D.; et al. Liquid Chromatography-Tandem Mass Spectrometry in Fat-Soluble Vitamin Deficiency. Clin. Chim. Acta 2023, 548, 117469. DOI: 10.1016/j.cca.2023.117469