A reminder of the continued relevance of macroenzyme detection testing

A reminder of the continued relevance of macroenzyme detection testing

Podsjetnik na kontinuiranu relevantnost testiranja na prisutnost makroenzima

Daniel Victor Šimac¹, Maja Šimac², Srđan Novak¹

¹Division of Rheumatology and Clinical Immunology, Department of Internal Medicine, University Hospital Centre Rijeka, Tome Strižića 3, 51000 Rijeka, Croatia / Zavod za reumatologiju i kliničku imunologiju, Klinika za internu medicinu, Klinički bolnički centar Rijeka, Tome Strižića 3, 51000 Rijeka, Hrvatska

²Department of Laboratory Diagnostics, University Hospital Centre Rijeka, Tome Strižića 3, 51000 Rijeka, Croatia / Klinički zavod za laboratorijsku dijagnostiku, Klinički bolnički centar Rijeka, Tome Strižića 3, 51000 Rijeka, Hrvatska

³Department of Internal Medicine, School of Medicine, University of Rijeka, Braće Branchetta 20, 51000 Rijeka, Croatia / Klinika za internu medicinu, Medicinski fakultet, Sveučilište u Rijeci, Braće Branchetta 20, 51000 Rijeka, Hrvatska

Corresponding author / Adresa autora za dopisivanje:

Daniel Victor Šimac

Division of Rheumatology and Clinical Immunology / Zavod za reumatologiju i kliničku imunologiju

Department of Internal Medicine / Klinika za internu medicinu

University Hospital Centre Rijeka / Klinički bolnički centar Rijeka

Tome Strižića 3, 51000 Rijeka

Croatia / Hrvatska

E-mail / e-pošta: danielsimac@hotmail.com Received / Primljeno: September 20, 2025 / 20. rujna 2025.

https://orcid.org/0000-0003-3821-6969Accepted / Prihvaćeno: December 2, 2025 / 2. prosinca 2025.

Dear Editor,

Although they were much more of a ‘hot’ topic back in the 1980s, and they are somewhat forgotten nowadays, macroenzymes should still be on the ‘radar’ of physicians who are working with patients. By their definition, macroenzymes are enzymes, such as transaminase (ALT, AST), amylase, lipase, and even creatinine kinase (CK). These enzymes are bound by immunoglobulins for the most part or polysaccharides in some cases, thus causing falsely elevated laboratory test results since these complexes are not effectively excreted by the kidneys. (1–3) They are generally considered benign, and they often remain undiscovered since they are commonly overlooked, but their relevance prevails. (1–4) The main issue is with physicians who are ordering additional, unnecessary, and sometimes invasive tests for further workup, or modifying effective treatment. This problem, when it is not recognized, actively burdens the healthcare system as well as the patients. (1–3) We have witnessed a handful of such cases firsthand, and one recent case comes to mind when we are discussing this topic. That is the case of a young man who underwent a number of tests before his diagnosis of macro-creatine kinase was confirmed, and this specific case serves as an excellent example of the aforementioned issue.

The case subject is a young male patient, 27 years old at the time, who was referred to a rheumatologist due to elevated CK levels. During a routine check-up, slightly elevated AST and ALT levels (115 U/L and 91 U/L) were found, and this finding was later supplemented with a finding of significantly elevated CK levels (8267 U/L). As the patient is physically active and exercises regularly, his CK levels were re-tested after a period of rest, and they were significantly elevated (3626 U/L). The patient did not report any signs of muscle weakness, skin rash, or other symptoms. Liver enzymes were found to be elevated before, approximately two years ago, and at that time serological testing for hepatotropic viruses and abdominal ultrasound were done. The results showed no evidence of abnormalities. At the time of referral, about a year had passed from a partial thyroidectomy, and the patient has been regularly taking a levothyroxine replacement drug. During further follow-up and workup, the patient remained asymptomatic, his CK levels were stable, and the ordered test results showed no abnormalities: myositis autoantibodies were tested and the result was negative, alpha-glucosidase activity result (to exclude Pompe disease) was unremarkable, and electromyoneurography (EMNG) findings did not show any signs of myopathy. The patient was examined by a neurologist, and cervical spine magnetic resonance imaging (MRI) was ordered. The results came back as normal. In the end, the presence of macroenzymes was considered, and testing was performed. The results were indicative of macroenzyme presence, thus confirming a final diagnosis of macro-creatinine kinase which served as an explanation for elevated CK levels. Perhaps elevated AST and ALT levels could be explained by macroenzymes as well, but tests were not performed to confirm this theory.

Macroenzymes can be found in healthy individuals, and they have been described in other cases (even iatrogenic, infective, and post-traumatic), but they are also associated with an autoimmune and lymphoproliferative disease, with the implication of polyclonal gammopathy, which is just one of the explanations for this phenomenon. (1–3) Interestingly enough, polyclonal gammopathy which was found in some cases of liver disease has also been shown to cause macroenzyme development. (2)

There are several methods which can be used to indirectly determine the presence of macroenzymes. One of them is polyethylene glycol (PEG) precipitation, a relatively simple and accessible test, already used in many laboratories for macroprolactin testing, which is a common laboratory procedure. (5) In short, the CK level is measured, and then PEG is applied which causes macroenzymes to precipitate, and following that the CK level is measured again. If there is a significant decrease between the measurements, this is considered as confirmation of macroenzyme presence. (5)

It is important to reiterate the importance of macroenzymes, but it is also important to note that, besides causing falsely elevated test results and confusion, some macroenzyme-related results may lead doctors to order more unnecessary tests, thus causing additional strain on the healthcare system, cost and patients. (1–4) One extreme example comes to mind in which the case subject was a patient with elevated AST levels, who ultimately underwent a liver biopsy procedure, a test which is quite invasive and carries certain risks. Following a liver biopsy, the diagnostic tests for macro-AST were performed and a macro-AST diagnosis was confirmed. (4) Another potential issue is the modification of effective treatment, during which the patient’s condition remained stable, due to mistaking macroenzymes for iatrogenic toxicity, which can possibly lead to the selection of alternative and less effective treatment options and disease exacerbation. (1–3)

Although it would not be prudent to only test for macroenzymes, in most applicable cases their consideration is certainly warranted, especially in seemingly healthy individuals, and potentially in other unclear cases. Although the popularity of macroenzymes as a topic has waned over the years, they continue to be relevant, and any practicing physician should be informed about this laboratory anomaly when interpreting test results.

Author Contributions: Article conception and design: DVS, MS, SN. Data acquisition, analysis and interpretation: DVS, MS, SSN. Article drafting: DVS, MS, SN. Critical revision of important intellectual content: DVS, MS, SN.

Acknowledgments: The authors report no acknowledgments.

Funding: For this work authors did not receive any funding.

Conflict of interest statement: The authors declare that they have no conflict of interest relevant to this manuscript.

REFERENCES / Literatura

1. Remaley ATR, Wilding P. Macroenzymes: Biochemical characterization, clinical significance, and laboratory detection. Clin Chem. 1989;40:2261.

2. Klonoff DC. Macroamylasemia and other immunoglobulin-complexed enzyme disorders. West J Med. 1980;133(5):392–407.

3. Šimac DV, Špelić M, Devčić B, Rački S. Diagnosing macroamylasemia in unexplained hyperamylasemia. Acta medica Croatica. 2017:63–66.

4. Briani C, Zaninotto M, Forni M, Burra P. Macroenzymes: Too often overlooked. J Hepatol. 2003;38:119.

5. Davidson DF, Watson Ann DJM. Macroenzyme detection by polyethylene glycol precipitation. Clin Biochem. 2003;40:514–520.