Copper and Caeruloplasmin

Copper is an essential trace element in cytochrome oxidase, superoxide dismutase, lysyl oxidase, dopamine beta hydroxylase and the ferroxidase caeruloplasmin. About 95% of serum copper is bound to caeruloplasmin, a copper carrying alpha-2 glycoprotein made in the liver, and the rest is loosely bound to albumin and amino acids. The liver removes excess copper into bile through ATP7B, and disorders of ATP7B and ATP7A cause Wilson disease and Menkes disease.

Serum copper and caeruloplasmin are measured by atomic absorption or inductively coupled plasma mass spectrometry (copper) and by immunonephelometry or an oxidase activity assay (caeruloplasmin). Twenty four hour urinary copper and calculated non-caeruloplasmin-bound copper complete the assessment.

The main use is the diagnosis of Wilson disease, in which copper accumulates in the liver, brain, cornea and kidney. The other uses are the diagnosis of Menkes disease and of acquired copper deficiency.

Wilson disease has a prevalence of about 1 in 30 000 and a carrier frequency of about 1 in 90. It is treatable, and early treatment prevents liver failure and neurological disability. Acquired copper deficiency is rare but is recognised after bariatric surgery, with zinc excess, in malabsorption and with long term parenteral nutrition.

Indications. Unexplained liver disease, acute liver failure with haemolysis, hepatitis or cirrhosis in a person under 40 years, movement disorder, psychiatric illness or cognitive change in a young person, Kayser-Fleischer rings, unexplained Coombs negative haemolytic anaemia, family history of Wilson disease, infants with hypotonia, seizures and kinky hair (Menkes), and anaemia or neutropenia with myelopathy (copper deficiency).

Specimen. Serum from a trace element tube (royal blue top) or acid washed plastic container, avoiding rubber stoppers that contain metals. Caeruloplasmin on serum. Twenty four hour urine in an acid washed, metal free container, with the collection complete (creatinine is checked). Haemolysis does not materially change serum copper.

Values are method specific. Conversion: copper microg/dL multiplied by 0.157 gives micromol/L, and caeruloplasmin mg/dL multiplied by 0.01 gives g/L.

Serum copper. Range: 11 to 22 micromol/L (70 to 140 microg/dL). Raised: pregnancy, oestrogen and oral contraceptive use, inflammation, infection, malignancy (lymphoma), biliary obstruction, and thyrotoxicosis. Lowered: Wilson disease (most patients), Menkes disease, nephrotic syndrome, protein losing enteropathy, malnutrition, malabsorption, parenteral nutrition without copper, zinc excess, and aceruloplasminaemia.

Serum copper is often low in Wilson disease because caeruloplasmin is low, and may be normal or high in acute liver failure when copper is released from the liver.

Caeruloplasmin. Range: 0.20 to 0.50 g/L (20 to 50 mg/dL). It is an acute phase protein. Raised: pregnancy, oestrogen, inflammation, infection and malignancy. Lowered: Wilson disease (typically under 0.10 g/L), Menkes disease, heterozygous carriers (often 0.10 to 0.20 g/L), aceruloplasminaemia (undetectable, with iron overload, diabetes and neurological disease), advanced liver disease, nephrotic syndrome, protein losing enteropathy, malnutrition, and copper deficiency.

A value under 0.20 g/L raises the possibility of Wilson disease. A normal caeruloplasmin occurs in 5 to 15% of patients with Wilson disease, particularly with inflammation, so a normal value does not exclude it, and about 20% of patients have a level over 0.20 g/L.

Non-caeruloplasmin-bound copper. Calculated as total copper (microg/dL) minus 3.15 times caeruloplasmin (mg/dL). Normal range: 5 to 15 microg/dL (0.8 to 2.4 micromol/L). Over 25 microg/dL (4 micromol/L) supports Wilson disease. The calculation is inaccurate with low caeruloplasmin values and with immunoassays that detect inactive protein.

24-hour urinary copper. Normal: under 0.6 micromol/24 hours (under 40 microg/24 hours). A value over 1.6 micromol/24 hours (over 100 microg/24 hours) supports symptomatic Wilson disease, and values of 0.6 to 1.6 micromol/24 hours need further tests. Urinary copper rises in cholestasis and nephrotic syndrome.

Liver copper. Hepatic copper over 4 micromol/g dry weight (over 250 microg/g) on biopsy is the best single biochemical test, though it can be lower with patchy cirrhosis.

Wilson disease.

• Presentation: hepatic (raised aminotransferases, steatosis, hepatitis, cirrhosis, acute liver failure) in children and young adults aged 5 to 35, neurological (tremor, dystonia, dysarthria, parkinsonism, ataxia) usually in the second to third decade, psychiatric (depression, personality change), and haemolytic.
• Diagnostic features: low caeruloplasmin, raised 24-hour urinary copper, Kayser-Fleischer rings on slit lamp examination (present in almost all with neurological disease and about half with isolated liver disease), raised hepatic copper, and biallelic pathogenic ATP7B variants.
• Acute liver failure in Wilson disease: Coombs negative haemolytic anaemia, very high serum and urinary copper, low alkaline phosphatase (ratio of alkaline phosphatase to bilirubin under 4), and AST to ALT ratio over 2.2.

Treatment monitoring. Urinary copper 200 to 500 microg/24 hours (3 to 8 micromol/24 hours) on chelation indicates adequate treatment, and under 75 microg/24 hours suggests overtreatment or poor adherence. Non-caeruloplasmin-bound copper of 5 to 15 microg/dL is the goal. With zinc, urinary copper is under 125 microg/24 hours.

Menkes disease. X-linked ATP7A disorder in infants: low serum copper and caeruloplasmin, hypotonia, seizures, kinky hair, failure to thrive, arterial tortuosity and bladder diverticula.

Acquired copper deficiency. Low copper and caeruloplasmin with anaemia and neutropenia (sometimes pancytopenia, copper deficiency mimics myelodysplasia), sensory ataxia with myelopathy, and optic neuropathy. Causes include zinc excess, gastric surgery, malabsorption, parenteral nutrition, enteral feeding, and excess antacid.

Aceruloplasminaemia. Neurological disease, diabetes and retinal degeneration with iron accumulation and absent caeruloplasmin, but low serum iron and high ferritin.

Critical values and findings needing immediate action.

• Acute liver failure or severe hepatitis with haemolysis in a young patient, which needs urgent exclusion of Wilson disease and transplant assessment.
• Serum copper over 40 micromol/L or urinary copper far over range with haemolysis.
• Anaemia and neutropenia in a patient with a low copper and ataxia.
• Infant with seizures and low copper.

Interferences and pitfalls.

• No single test excludes Wilson disease, and several are combined.
• Caeruloplasmin is raised by pregnancy, oestrogen, inflammation and infection, giving false normal results.
• Caeruloplasmin is low in other chronic liver disease, malnutrition, nephrotic syndrome and in carriers, giving false positives.
• Immunoassays for caeruloplasmin measure inactive protein too, and enzyme activity assays are more accurate in some cases.
• Urine collections contaminated with metal or incomplete give wrong values.
• Chelator therapy increases urinary copper and cannot be used for diagnosis.
• The Kayser-Fleischer ring needs a slit lamp, and may be absent in young patients.
• Siblings of a patient need testing even if asymptomatic.

Special groups.

• Neonates and infants: caeruloplasmin is low in the first 6 months and rises to adult levels at about 1 year, so age specific ranges are needed.
• Children: Wilson disease can present with isolated liver disease from age 3 years.
• Pregnancy: copper and caeruloplasmin rise by 2 to 3 times, and treatment of Wilson disease is continued with dose adjustment.
• Older adults: copper deficiency should be considered in unexplained cytopenias and myeloneuropathy.
• Regions: consanguinity increases frequency of Wilson disease, and Indian childhood cirrhosis and idiopathic copper toxicosis are associated with copper containing utensils.

Follow up tests. ATP7B gene sequencing, slit lamp examination for Kayser-Fleischer rings, liver function tests and INR, full blood count and film, direct antiglobulin test, liver biopsy with copper quantification, abdominal ultrasound, MRI of the brain, urine copper before and after penicillamine, family screening of siblings, zinc level, iron studies and ferritin, and copper level in liver tissue.