Vitamin Tests

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Vitamin Tests


Vitamins are small organic molecules that are necessary for many biochemical reactions in the body. For example, many vitamins participate as cofactors in enzyme reactions within the cells. They must be obtained through diet, microorganisms in the gut or sunlight since humans cannot synthesize them. Vitamin tests measure the levels of certain vitamins in an individual's blood which can be correlated to the levels of vitamin in their tissues. They are generally used to aid in the diagnosis of vitamin deficiencies or in detecting toxic amounts of a vitamin in a patient's system.


Vitamins are components of food that are needed for growth, reproduction, wound healing, digestion of food, blood clotting, bone metabolism and maintaining good health. They can be isolated from plants and organisms or they can be synthesized in the laboratory. The vitamins include vitamin D, vitamin E, vitamin A, and vitamin K, which are the fat-soluble vitamins, and folate, vitamin B12, biotin (vitamin H), vitamin B6, niacin, thiamine, riboflavin, pantothenic acid, and ascorbic acid (vitamin C ), which are the water-soluble vitamins. Fat soluble vitamins are absorbed and transported differently from the water soluble vitamins. They can remain in the body longer since they can be stored in fat so they are more toxic in high doses.

Vitamins are required in the diet in only tiny amounts, in contrast to sugars, starches, proteins and fats. However, vitamin requirements can rise after surgery, with cancer and other illnesses, and during infection and pregnancy. Not receiving sufficient quantities of a certain vitamin can be devastating, resulting in vitamin deficiency diseases such as scurvy (vitamin C deficiency), pellagra (niacin deficiency), megaloblastic anemia (vitamin B12 or folate deficiency) or rickets (vitamin D deficiency). Less extreme deficiencies can cause a delay in wound healing. Conversely, consuming too much of a certain vitamin, especially the fat soluble ones, can be toxic to a person's system. While most vitamin deficiencies are rare in our society, they can be caused by certain diseases, especially those that affect absorption of food, or can be caused by inborn errors of metabolism, fad diets, anorexia, blood loss, parenteral nutrition and dialysis. The vitamins that are most commonly measured by doctors are folate, vitamin B12, vitamin K, vitamin D, and vitamin A.


Most vitamin tests are performed on blood samples collected from a vein in the crease of the arm. The nurse or phlebotomist performing the procedure should observe universal precautions for the prevention of transmission of bloodborne pathogens. Some drugs are known to increase or decrease the level of specific vitamins. The physician should obtain a thorough list of the patient's medications when requesting vitamin measurements.


Many of the vitamin tests done today are vitamin status panels, such as megaloblastic anemia panels that measure the concentration of both vitamin B12 and folate. A deficiency of either of these vitamins results in anemia associated with enlarged (macrocytic) red blood cells. The actual testing methods take advantage of the compound's chemical composition. Fat soluble vitamins are measured differently from water soluble vitamins. In general, the tests are performed on plasma, although some tests for metabolites of vitamins can be done on urine, as is the case with many of the water soluble vitamins. Each vitamin occurs at extremely small concentrations in the blood and urine when compared to levels of most other molecules. For this reason, a procedure that separates the vitamin from the rest of the compounds in the sample is usually performed immediately prior to conducting the actual test. This isolation is done using filters that allow the vitamin to pass through and leave the bigger molecules behind. Vitamin B12 and folate are routinely measured by immunoassay methods, but the most common method to measure other vitamins is high-performance liquid chromatography (HPLC). In HPLC, the vitamin to be measured is extracted from the sample and injected into a stream of solvent that is pumped at high pressure through a column packed with particles to which an organic liquid has been bonded. The molecules separate at different rates depending upon their affinity for the bonded particles. The time at which they elute (i.e., come out of the column) is used to identify the molecules. As the vitamins elute, they flow through a detection cell where they are measured by ultraviolet or infrared light absorption or by fluorescence. In these reactions, the amount of light absorbance or amount of fluorescence is proportional to the amount of vitamin in the sample.

While HPLC and immunoassay are the testing methods used most often, other types of tests exist including biochemical (photometric) tests and microbiological assays. Some tests, such as those for riboflavin, are conducted by giving the patient a riboflavin "load" and looking at metabolites in the urine. Vitamin K defiency, a vitamin crucial in blood clotting, is often evaluated by a surrogate test, the prothrombin time. The prothrombin time is measured routinely on patients before they undergo a surgical procedure since long clotting times can complicate surgeries. The test measures how long it takes for a fibrin clot to form in a plasma sample to which calcium and tissue thromboplastin (a clot activator) have been added. Clotting factors II, VII, XI, and X are made from vitamin K. The prothrombin time is prolonged when there is a deficiency of fibrinogen or factors II, V, VII, or X. Therefore, a prolonged prothrombin time can result from an inherited or acquired deficiency of one of these factors or a deficiency of vitamin K.

Other vitamins can be measured by taking advantage of their functions. One example is the measurement of oxidation/reduction reactions to measure vitamins E and C, two major antioxidants. In addition, many of the water soluble vitamins such as vitamin B6 are measured using microbiological tests. Samples, usually filtered urine, are applied to a culture medium. The medium is inoculated with a standardized concentration of the bacteria or yeast that requires the vitamin for growth. The plate is incubated for several days to see if the organism grows. Lack of growth indicates a low concentration of the vitamin and correlates well with vitamin deficiency.


Most vitamin tests require no preparation; however, some may require that the patient fast for at least eight hours before giving a blood sample, or stop using some medications.


Levels of vitamins in the body must be interpreted carefully. Many times low levels do not correlate with disease and the patient is asymptomatic. Other times the levels may seem fine, but the patient displays symptoms of a deficiency. Physicians must take into account the patient's dietary history (e.g., is the patient a vegetarian?), medications, and also do a thorough physical exam.


Antioxidant— A compound that protects against oxidation, usually by being oxidized itself. Antioxidant vitamins include C and E.

Carotenoids— Pigments found in vegetables and fruits, similar in structure to vitamin A. These compounds act as antioxidants.

Megaloblastic anemia— A disorder caused by a deficiency in vitamin B12 and folate. The blood contains immature red blood cells causing a decrease in oxygen carrying capacity and symptoms of fatigue and peripheral neuropathies.

Parenteral nutrition— Also called total parenteral nutrition or TPN. A slang term is "tube feeds." Parenteral nutrition is the taking in of nutrients through any way other than by the gastrointestinal system. TPN can be administered by intravenous, subcutaneous, intramuscular or intramedullary injection. TPN formulations need to be prescribed based on the patient's illness and how long they will be on the feeding regimen.

Pellagra— A disorder caused by a deficiency in niacin. The patient will have dementia, diarrhea and dermatitis.

Peripheral neuropathy— A disorder characterized by tingling and numbness in extremities. This can cause difficulties with walking and using your hands.

RDA— Recommended dietary allowance. The amount of vitamin needed to maintain a healthy level in tissues by a healthy person.

Rickets— A disorder caused by a deficiency in vitamin D. The symptoms are muscle hypotonia (weak muscles) and skeletal deformity. Seen mainly in children.

Scurvy— A disorder caused by a deficiency in ascorbic acid, or vitamin C. The patient will have swollen and bleeding gums, loss of teeth, skin lesions and pain and weakness in lower extremities.

Vitamins— Small compounds required for metabolism that must be supplied by diet, microorganisms in the gut (vitamin K) or sunlight (UV light converts pre-vitamin D to vitamin D).

Representative normal ranges for certain vitamins are listed below. Normal ranges vary depending upon the patient's age and method of analysis. Levels of some vitamins will be different in pregnancy. Please note that, by convention, the units used for reporting one vitamin may differ from another. The units picogram/milliliter (pg/mL), nanogram/milliliter (ng/mL), and micrograms per deciliter (micrograms/dL) refer to the weight of vitamin in the specified volume. The units nanomoles/liter (nmol/L) and micromoles/liter (umol/L) refer to the concentration of vitamin in the specified volume.

  • folate (folic acid ): 3.0-20.0 ng/mL in serum, 140-628 ng/ml in red blood cells
  • vitamin B12: 200-835 pg/mL
  • thiamine: 9-44 nmol/L
  • riboflavin: 4-24 micrograms/dL
  • vitamin B6: 5-30 ng/mL
  • vitamin C (ascorbic acid): 0.4-1.5 mg/dL
  • vitamin A: 30-80 micrograms/dL
  • vitamin D (25-hydroxy-vitamin D): 14-60 ng/mL
  • vitamin K: 13-1190 pg/mL
  • vitamin E: 0.5-1.8 mg/dL

Health care team roles

Tests for specific vitamins are requested by a physician. Vitamin deficiencies may be suspected by physicians and nurses, and patients who are deficient should be referred to a dietician who can advise them on food choices. In addition, health care providers should be vigilant for hidden deficiencies in those who are pregnant, have malabsorption disorders or chronic illnesses such as cystic fibrosis. Blood samples are collected by a nurse or phlebotomist. Vitamin assays are performed by clinical laboratory scientists/medical technologists.



Burtis, Carl A., and Edward R. Ashwood. Tietz Textbook of Clinical Chemistry. Philadelphia: W.B. Saunders Company, 1992.

Kaplan, Lawrence A., and Amadeo J. Pesce. Clinical Chemistry Theory Analysis and Correlation. St. Louis: Mosby Publishers, 1996.