Introduction

Diabetes mellitus (DM), a global epidemic, is increasing at an alarming rate and is associated with both increased morbidity and mortality. In 2018 the estimated prevalence of diabetes in the USA was 34.2 million people. Globally, the prevalence was 425 million people and this is expected to rise to 629 million by 2045. Currently, only plasma glucose and glycated hemoglobin (HbA1c) are universally accepted as reliable measures of diabetes control. In certain conditions, the HbA1c measurement is not reliable. An example is in patients with red blood cell (RBC) disorders and renal disease. Fructosamine, which is a measure of non-enzymatic glycation of circulating proteins including albumin, globulins, and lipoproteins, has evolved to be a  reasonable alternative to HbA1c measurement in situations where HbA1c is not reliable. Because albumin is the most abundant of the serum proteins, fructosamine is predominantly a measure of glycated albumin (GA), which represents the percent of albumin that is glycated. Fructosamine and GA have a potential role in the diagnosis, monitoring, and management of diabetes. [1][2][3]

Pathophysiology

HbA1c is a product of non-enzymatic glycation of hemoglobin. Red blood cells have a lifespan of approximately 90 to 120 days, hence HbA1c indicates the mean blood glucose concentration over the lifespan of the RBC. HbA1c is influenced by conditions affecting RBC survival. Conditions causing low RBC turnover like untreated iron, vitamin B12, or folic acid deficiency anemias, will result in falsely high HbA1c values. On the other hand, conditions causing high RBC turnover like hemolytic anemia, in patients treated for iron, vitamin B12, or folate deficiencies, as well as in patients treated with erythropoietin (like in chronic kidney disease), there will be falsely low HbA1c values.

Fructosamine (1-amino-1-deoxy fructose), is a stable ketoamine, formed by the reaction between glucose and the amino group of protein (predominantly albumin, but also including globulins and lipoprotein). The attachment of the aldehyde group of the carbohydrate with the N terminal amino acid of the protein forms the reversible Schiff base product, the aldimine intermediate. The Schiff base product may be converted back to glucose and protein, or undergo the Amadori rearrangement to form stable fructosamine. This process is known as non-enzymatic glycation and is also referred to as the Maillard reaction. The Maillard reaction causes the browning phenomenon that occurs in milk and other food products when heated. Glycated albumin refers to the formation of ketoamine specifically involving the major circulating protein albumin (3.5 g/dl to 5 g/dl). Glycated albumin is an example of a fructosamine (FA). Because albumin is the most abundant of the serum proteins, fructosamine is predominantly a measure of glycated albumin. The formation of fructosamine and glycated albumin are post-translational modifications that occur to proteins.

Non-immunoglobulin serum proteins have a much lower half-life, approximately 14-21 days.[4] The measurement of fructosamine or GA provides information on glucose control within the previous 2-3 weeks. Another important difference with HbA1c is the rate of nonenzymatic glycation of albumin, which is approximately 9- to 10-fold higher than that of HbA1c.[5][6]

Specimen Requirements and Procedure

Sample type: Serum or plasma are the sample types used for the measurement of fructosamine and glycated albumin. Fasting specimens are not required.

Testing Procedures

Fructosamine

The most common assay available for fructosamine measurement in serum is the colorimetric-based assay. This assay utilizes the reduction of the dye nitroblue tetrazolium (NBT) to formazan. The rate of formazan formation is directly proportional to the fructosamine concentration and is measured with the spectrophotometric technique.[7] These assays are widely available, can be automated, and fairly inexpensive. The reference range for fructosamine in non-diabetic individuals is generally 200 to 285 umol/L. However, unlike HbA1c, there is a serious lack of standardization across the different fructosamine assays.

Glycated Albumin

There are several different assay methodologies available for the analysis of glycated albumin. These include:

• Enzymatic assay
• High-performance liquid chromatography (HPLC) and affinity chromatography
• Immunoassay, including quantification by radioimmunoassay
• Enzyme-linked immunosorbent assay (ELISA)
• Enzyme-linked boronate immunoassay (ELBIA)
• Colorimetry
• Electrochemical

The enzymatic assay (Lucica GA-L kit, Asahi Kasei Pharma, Tokyo, Japan) is easier to use, highly accurate, and automated.[8] First, there is the elimination of endogenous glycated amino acids and peroxide by a ketoamine oxidase, followed by a peroxidase reaction.[9] An albumin-specific proteinase hydrolyzes the GA. The products of this reaction are oxidized by ketoamine oxidase to hydrogen peroxide, which is then measured quantitatively by a colorimetric method. The albumin concentration is also measured concurrently. The final result is expressed as the ratio of glycated to total albumin.[10]

The normal value is around 14% and it becomes greater than 17% in diabetes patients. Values in diabetes can go as high as two to five times the upper limit of normal.

Interfering Factors

Fructosamine assays are affected by changes in temperature and by the increased presence of reducing substances in serum, for example, vitamin C and bilirubin. Fructosamine and GA both do not have standardized assays. Additionally, both fructosamine and glycated albumin are affected by the presence of any conditions that influence serum albumin concentrations. However, this is minimized for GA since this is expressed as a percentage of total albumin. Fructosamine will be unreliable when serum albumin is less than 3.0 g/dl. This will include conditions where there is decreased albumin synthesis, like in liver cirrhosis, or when there is albumin/protein loss such as in nephrotic syndrome and protein-losing enteropathies. Fructosamine levels may also be affected by conditions with raised total protein levels, like in multiple myeloma (due to increased immunoglobulins) and in polyclonal gammopathies.

Results, Reporting, and Critical Findings

A  reference range for fructosamine in non-diabetic individuals is generally 200 to 285 umol/L.

While GA assays also suffer from standardization, the newer assay developed by Asahi Kasei appears to be much improved. According to this assay, normal persons have values around 14% and those with diabetes greater than 17%. Values in diabetes can go as high as two to five times the upper limit of normal.

Clinical Significance

The clinical utility of fructosamine and GA includes monitoring of diabetes, diagnosis of pre-diabetes, and prediction of both the microvascular and macrovascular complications. They have the advantage of not requiring a fasting sample.

Monitoring of  Glucose Control in Diabetes

Fructosamine and glycated albumin can be utilized as short-term markers of glucose control.  Both correlate significantly with HBAIc levels. While HbA1c reflects glucose control over a period of the preceding 8 to 12 weeks, fructosamine reflects the average glycemia over the preceding 2 to 3 weeks. This is a result of the inherent shorter half-life of albumin in comparison to hemoglobin in the erythrocyte.

Fructosamine has largely been used as an alternative to the use of HbA1c monitoring in the presence of certain conditions that preclude the use of HbA1c, such as hemoglobin variants and alterations in erythrocyte lifespan. Fructosamine and glycated albumin are not affected by hemoglobin level, or red blood cell characteristics to which HbA1c is susceptible. This includes conditions such as hemoglobinopathies, sickle cell anemia, and anemia related to iron, vitamin B12, or folate deficiency.

Additionally, fructosamine has clinical utility in conditions where information regarding short-term glucose control is important in the management of the patient such as in pregnancy, or recent medication adjustment. FA and GA can also be useful in monitoring people with diabetes with fluctuating or poorly controlled diabetes.

Diagnosis of Diabetes

Recent studies have evaluated the use of the alternate glycaemic markers of fructosamine and glycated albumin for the diagnosis of diabetes. It has been reported that in the diagnosis of diabetes, serum GA measurements can be used to ascertain the need for an oral glucose tolerance test (OGTT). There appears to be a negative correlation between GA and body mass index (BMI), and hence it could potentially underestimate glycemia in the obese. Currently, no guidelines support the use of GA or FA for the diagnosis of diabetes or pre-diabetes.[10]

Diabetes Outcome

Previously there was little evidence of the relationship of fructosamine and glycated albumin with diabetes complications and long-term outcomes. Recent studies, like for example, the Atherosclerosis Risk in Communities Study (ARIC), have demonstrated that fructosamine and glycated albumin were strongly associated with retinopathy as well as significantly associated with the risk of incident chronic kidney disease and incident diabetes. Besides, both Fructosamine and GA, even following adjustment for HBAIc, are significant prognosticators of cardiovascular outcomes and mortality.[11]

Quality Control and Lab Safety

Commercial assays for fructosamine and glycated albumin have internal quality control materials available for use. Additionally, laboratories measuring these assays would subscribe to a recognized proficiency testing scheme to monitor test performance.

Enhancing Healthcare Team Outcomes

Healthcare workers including the nurse practitioner should be familiar with the diagnosis of diabetes. Fructosamine and GA can be utilized as alternate markers in those patients where the HbA1c assay is unreliable. Also, they can identify poor glucose control more rapidly than HbA1c, i.e., short-term hyperglycemia. A major promise of the tests is their ability to predict those pre-diabetic patients who progress to clinical diabetes since this could lead to major lifestyle and pharmacological interventions to prevent the onset of diabetes and its complications. Finally, they may also have a role in the management of diabetes during pregnancy since pregnant patients need frequent glucose monitoring. They can provide a measure of glycemia over 2 to 3 weeks rather than 8 to 12 weeks as is with HbA1c.[12]

Glycated albumin has been reported to be a better marker than HbA1c  for the assessment of glucose control in people with diabetes with chronic kidney disease and those on hemodialysis and peritoneal dialysis.[13][14]