Preface

When an unexpected laboratory test result occurs in the absenceof an in other respects obvious etiology, a useful rule is toconsider a drug effect. Many drugs affect the outcome of laboratory tests. Knowledge about whichdrugs cause erroneous results is essential to avoid misleadingconclusions as well as unnecessary expenses for medical care.The importance of interference with clinical laboratory analyseswas reviewed by M.H. Kroll and R.J. Elin in Clinical Chemistry1994;40: 1996-2005.

Drugs can influence biochemical and hematological investigationsin two ways:

A. - In vitro, i.e. interference with the analyticalprocess.

B. - In vivo, i.e. biological effects, which may beeither an effect of the main action of the drug or a side effect.

A. Analytical interference of drugs: in vitro effects.

A drug or its metabolites may interfere with the analytical techniquesin several ways. Drugs which interfere with one analytical techniquewill not necessarily interfere with other methods and thereforethe specific features of each analytical method should be known.Slight differences in technique may be decisive for the degreeof interference caused by the drug. This applies to the determinationof creatinine, which may be affected by drugs as ascorbic acidand methyldopa. Ascorbic acid is widely used. Large doses of vitaminC can influence several analytical techniques.

Ascorbic acid is a common source of interactions in the estimationof glucose, hemoglobin and nitrite in urine. Even if many modernreagent sets claim insensitivity high doses of ascorbic acid frequentlyused as dietary supplement can still interfere. Therefore, patientsshould avoid consumption of high doses of ascorbic acid 24 hoursbefore urine collection for routine examinations.

In addition ascorbic acid can affect the analysis of blood infeces by giving a false negative result and thus the diagnosisof a bleeding state might be overlooked. Iron salts have earlierbeen reported to cause false positive results with some commonlyused tests for fecal hemoglobin. However, according to recentpublications this is not clinically relevant. Consequently, itis necessary to follow and reevaluate the information.

The analytical methods have been substantially developed and modifiedin recent years and more specific and reliable methods are nowavailable. However, even specific methods may be liable to interferencesand this applies to colorimetric and enzymatic methods as wellas methods based on immunological principles.

Paracetamol can interfere with the blood glucose determinationusing the YSI glucose analyzer. One mmol of paracetamol per litreincreases the apparent glucose concentration by 3-5 mmol/L. Thefalse increase in glucose values can be of considerable importancein patients with high paracetamol dosages. The manufacturer ofthe instrument has produced a modified enzyme membrane which preventsthe interference. However, the result can vary depending on thestatus of the membrane. In addition the old type of techniqueis still for sale. Another analytical drug interference is thedepression of serum creatinine values in creatinine-PAP-methodsby calcium dobesilate.

Instruments for use nearer the patient have appeared in recentyears, i.e. Seralyzer, Reflotron and Ektachem DT 60. These applystrictly standardized techniques but are not entirely free fromdrug interferences. A comprehensive book covering dry chemistryanalyses has been published by O. Sonntag (Dry Chemistry, ElsevierScience Publishers, Amsterdam, 1993). The analytical interferenceswhich have been evaluated as clinically relevant have also beenincluded in the present lists.

Infusion of fat emulsions may disturb the electronic countingof erythrocytes and leucocytes as well as certain spectrophotometricalanalyses. Laboratory results can therefore be greatly misleadingif blood samples are taken during or too close after an infusion. The interferences in commonly available analyticalsystems from lipemia, including the effect of fat emulsions, havebeen extensively evaluated by M.R. Glick and K.W. Ryder (ClinChem 1987;33:1453-1458). This literature is also relevantin out-patients due to the fact that blood specimens are oftentaken from non-fasting persons.

In a review "Interferences due to lipaemia in routine photometricanalysis - survey of an underrated problem" by J.Brady and N. OíLeary in Ann Clin Biochem 1994;31:281-288the authors summarise that many analyses are affected by lipemiaand that the degree of interference is instrument/method dependent.

Ideally, information on administered drugs and their possibleeffects on analytical performance should accompany all test requestsand reports respectively. Obviously this requires a sophisticatedcomputer system which is rarely available today. The present compilationis an attempt to be one source of information.

Studies to evaluate the drug interference in clinical chemicalmethods

When a new analyzer or new method is introduced into laboratorymedicine an interference study is a major part of an evaluationreport. You may find information in the literature or given bythe supplier or manufacturer of the instrument or reagent. Forthose who want to make their own interference study a proposalis given in the following chapter.

Two different types of studies are needed to characterize theexistence or degree of interference by drugs. The first studyinvolves estimating possible interference by drugs within a concentrationrange which is higher than the therapeutic concentration of therelevant drug. The second type of study involves only those drugswhich have caused an interference in the first type of study toshow the magnitude and the relationship between the concentrationof the analyte and the degree of interference.

To select the suitable concentration of the drug in the firsttype of study it is necessary to be aware of the meaningful concentrationwhich is not above the lethal concentration of the respectivedrug. As a rule of thumb the maximal therapeutic concentrationshould be multiplied by five to reach the test concentration.This procedure has some limits because there is no direct correlationbetween the therapeutic concentration and the lethal or toxicconcentration. The factor is often higher or lower than 5. Therefore,when the toxic, lethal or peak concentrations are known theseshould be used instead of the theoretical value mentioned above.

Selection of drugs for the first type of study
The selection of the drugs described in Table 1 is based on alarge literature search. The listed drugs are known to cause interferencesin some analytical methods. The table is not a definitive listof the drugs most commonly used. In some studies it is necessaryto use a broader spectrum of drugs or alternative drugs to checkthe interference in a special area of interest. Some alternativedrugs which are often used are therefore listed in Table 1 too.To interpret the clinical significance of an observed drug interferenceand to reduce the problems of erroneous laboratory results additionalpharmacokinetic data are given in Table 1 (see also "Interpretationof a suspected interference").

A drug can interfere with clinical chemical investigations inseveral ways: By direct interference with the chemistry of thedetermination, causing an increase or a decrease in the detectedreaction product or by inhibiting an enzyme or an antibody reactionthat is part of the analytical procedure. Evaluation experimentsare needed to obtain information on the presence and the extentof the drug interference. Some drugs have strong biochemical effectsand must always be suspected to cause analytical interferences.At a meeting in Penzberg, Germany in December 1995 with expertsfrom seven different European countries the following 18 drugswere selected because they may cause interferences on serum orplasma determinations.

Criteria used for selection were:

The following drugs were chosen:

Selected candidates: Additional candidates:
Acetaminophen (paracetamol) N-Acetylcysteine
Acetylsalicylic acid Ampicillin
Ascorbic acid Calciumdobesilate
Heparin Na2-cefoxitin
Intralipid (fat emulsions) Ciclosporine
Levodopa Ibuprofen
Methyldopa Metronidazole
Theophylline Phenylbutazone
Rifampicin
Tetracyclin

Details will be published by the organizer of the meeting ProfessorBreuer, Gelsenkirchen.

Procedure of the drug interference studies

Pooled serum or plasma
Drug free serum or plasma from healthy adults is pooled to beused in the interference studies. Usually concentrations aroundthe decision limit of the analyte should be used. For some analytesit is useful to have samples with higher concentrations as recommendedbecause of the poor analytical precision at the lower range. Concentrationsor activities close to the detection limit of the method in useshould be avoided.

Drug concentrations
The toxic or lethal concentration of the drug should be used ifavailable. If the toxic or lethal concentration is not known theconcentration used in the study should be 5 times higher thanthe stated typical concentration in Table 1.

Solubility
Since not all drugs or their salts are soluble in water, plasmaor serum different solvents must be used sometimes. If anothersolvent is used the effect of the solvent on the studied methodmust be checked prior to the start of the study.

The following solvents should be used (Table 1):1: pooled serum or plasma2: 0.1 ml 0.25 n NaOH and 9.9 ml pooled serum or plasma3: 0.1 ml 70% ethanol and 9.9 ml pooled serum or plasma

The solvent described above is used also as the blank sample ifno drug is added. The required amount of drug (see "Drugconcentrations") for the study should be weighed with abalance and dissolved in a separate volume of solvent 1 - 3.

Stability
After preparation of the sample containing the substances whichmay cause an interference the measurement of the analyte shouldbe done within a short period of time. Interactions or reactionsof the drug with sample constituents are also possible. The timebetween the preparation and measurement should not be longer than2 h because of the instability of the drug in the sample.

Measurement sequence
To evaluate the interference the following measurement sequenceis recommended (in brackets the number of measurements):

A. Calibration of the method or system according to the recommendationof the manufacturer
B. Measurement of the quality control materials (low and high)
C. Pooled serum or plasma without the added drug (blank sample)(n = 5)
D. Five different pooled sera or plasmas with added drug (n = 2)
E. Pooled serum or plasma without added drug (blank sample) (n = 5)
F. Five different pooled sera or plasmas with added drug (n = 2)
and so on.

The sequence should end with the material without added drug (blanksample) and the quality control samples to confirm the stabilityof the analytical system (reagents and instrument). Dependingon the number of samples to be measured quality control sampleshave to be analyzed more often (refer to instrument, regulatoryor other valid recommendations).

Statistical estimation and the second study
The mean value x and the standard deviation s of all data of theblank sample are calculated. Values of the sample with added drugsoutside the x +/- 3s range should be stated as a possible interference.

In this case the second study for assessing the amount of interferenceshould be prepared with different drug and analyte concentrations.For this study 5 different concentrations of the analyte coveringthe linear measurement range and 5 different concentrations ofthe drug covering the therapeutic and toxic range should be analysedas described above. For the presentation of these results a three-dimensionalplot according to Kroll, M. et al (Clin. Chem. 1987; 33:686-745)should be used. To better understand and describe the interferenceand its clinical significance the therapeutic and or the maximal(peak) concentration should be stated. The relationship betweenthe amount of interference, the drug concentration and the analyteconcentration may be linear or non-linear. The interference maybe different in various concentration ranges of both the drugand analyte.

Preparation of an interferogram
If only different concentrations of a drug were used to describethe interference on a single concentration of the analyte an interferogramcan be prepared according to Glick, M. R. et al (Clin. Chem. 1983;29:1208,Abstr. 389). The results obtained should be plotted on a x-y plotafter calculation of the recovery according to the formula

mean value (sample with addition of drug) x 100 /
mean value (blank samples)
= recovery (%)

The studied concentrations of the drug are drawn on the x-axis.The calculated recovery of the analyte is drawn on the y-axis.At the starting point (no drug added) the recovery is 100 %. Adeviation of +/- 10 % (or regarding the laboratory requirements)is allowed and should be drawn on the diagram. Every recoveryof the analyte and the remaining drug concentration are registeredas a point in the diagram. Beginning from the starting point eachpoint is linked with the next to produce an interference curve.The therapeutic concentration range if known is marked with twoarrows. An example is shown in Figure 1. If the plotted curvepasses under or over the limit this is considered to be an interference.The statement is valid only for the studied concentration of theanalyte. For other concentrations of the analyte different amountsof interference may occur. In these cases a three-dimensionalplot is needed to show the effect as mentioned above.


Figure 1: Interferogram. (Note: Arrows represent the therapeuticrange.)

Interpretation of a suspected interference
When a laboratory result is questioned by the clinical chemistor by the clinician detailed informations of the sample and thepatient have to be considered.
Is the sample coloured?
What is the diagnosis?
Which drugs have been taken by the patient?
At what time after the intake of the drug the sample was drawn?
Was the sample drawn out of an infusion line?
What privious laboratory results are available?

Sometimes the laboratory can give an explanation to the possibleinterference. It is important to know if the drug was taken shortlybefore the sample was drawn. Find out what t1/2 and at what timethe peak concentration of the drug appears (Table 1). It is necessaryto take a new sample before another application of the drug isgiven to the patient. Information regarding the actual interferenceis sometimes available - see lists in this book.

The laboratory method or technique must be well known. Some drugssuspected to interfere may be bound to proteins. The protein bindingvaries from drug to drug and in various clinical situations. Sometechnologies (e. g. ultrafiltration) eliminates the protein andtherefore also protein bound drugs and are sometimes suitablefor studies of interference.

B. Biological drug effects: in vivo effects

Biological effects of drugs which influence the interpretationof laboratory results may sometimes be found in all persons treatedwith a certain drug. Examples of such regular effects are seenduring hormone therapy. Although these effects are due to generalbiological and biochemical phenomena, they are included in thelists. The underlying mechanism is often discussed in the referencesincluded.

Sexual hormones often have considerable metabolic effects andcause significant changes in the plasma protein pattern. The plasmaconcentrations of orosomucoid, haptoglobin and albumin decreasewhile the concentrations of lipoproteins, transferrin, plasminogenand alpha-1-antitrypsin increase. The increases of ceruloplasmin,transcortin and thyroxine-binding globulin (TBG) are extremelyhigh. In women taking oral contraceptives or persons on estrogentherapy this is accompanied by an elevated concentration of serumthyroxine and cortisol. The free serum thyroxine level does not change, and estrogen therapy does notalter the thyroid gland function. However, from the elevated thyroxinevalues the physician may be misled to diagnose hyperthyroidism.

Oral contraceptives change lipid metabolism. Old types of contraceptivesraiseserum triglycerides as well as serum cholesterol. High densitylipoprotein cholesterol increases under the influence of the estrogen componentand decreases under the influence of the gestagen component. Theseeffects have become less prominent when the hormone content ofthe oral contraceptives has been reduced. It is sometimes no longerpossible to observe significant differences in concentrationsof serum lipids before and during low-dose treatment. The glucoseelimination rate may be lowered by traditional contraceptivesbut seems to remain uninfluenced by low-dose oral contraceptives.

The increase of amylase in serum or urine after administrationof morphine, codeine or pethidine may cause misinterpretation. Phenytoin andbarbiturates regularly increase serum gamma glutamyltransferaseby enzyme induction in the liver. Antiepileptics produce elevationof serum gamma glutamyltransferase in more than 80 per cent ofthe patients. Elevation of this enzyme is interpreted by manyclinicians to be an indicator of alcoholism. However, the factthat many drugs (including disulfiram) can cause the same effectmust be borne in mind.

Lithium carbonate therapy can be associated with myocardial damagecausing serum aminotransferase and creatine kinase elevations.

Intramuscular injections of drugs may sometimes be overlookedas a cause of increase in serum enzymes (e.g. creatine kinase).

Other examples of drug effects are hypercalcemia during treatmentwith thiazides and elevated serum creatinine in patients takinghigh doses of acetylsalicylic acid. The latter effect might bedue to suppressed tubular excretion of creatinine.

Interactions between drugs often influence the results of laboratorytests. The interactions include modifications of intestinal absorption, competitionin protein binding and transport of drugs. The effect on the metabolismof one drug by co-administration of another may be very important.

Antiepileptics accelerate the metabolism of many other drugs,for example anticoagulants. Many interactions have been reportedfor coumarin derivatives, resulting in either increased or decreasedcoagulation activity in plasma. It is likely that the importanceof the interactions varies from patient to patient. Certain drugse.g. cholestyramine can reduce the absorption of coumarin. Rifampicin,griseofulvin, barbiturates, glutethimide, phenazone and otherdrugs may weaken the effect of coumarin by accelerating coumarinmetabolism through enzyme induction. High doses of quinidine,clofibrate, sulfonamides and acetylsalicylic acid accentuate theanticoagulation effect by reducing the binding of coumarin toplasma proteins.

Many drugs affect the liver function and, accordingly, biochemicaltests monitoring the liver may become pathological. For reportsof liver injuries new methods for drug causality assessment havebeen published after international consensus meetings by G. Dananand Ch. Benichou in J Clin Epidemiol 1993;46:1323-1330.Often a certain pattern of analytical results evolves which isnot necessarily dose dependent. The abnormal test results arealmost always reversible after withdrawal of the drug. Individualpatients metabolize drugs differently which may cause variousand individual effects on biochemical markers.

Drugs may also be responsible for renal damage. Sulfonamides maycause different reactions e.g. glomerulonephritis and tubularnecrosis. Nephrotoxicity is often dose-related. Some drugs haveno effects on the kidneys of healthy individuals but may affectpatients with impaired renal function. Renal damage may resultfrom synergistic drug effects. Several drugs have been reportedto produce systemic lupus erythematosus (SLE) or alterations in laboratory tests indicatingSLE.

IFCC Committee on Drug Effects in Clinical Chemistry

The International Federation of Clinical Chemistry, IFCC has recognizedthat drugs can influence the interpretation of results from laboratoryinvestigations. - An Expert Panel on Drug Effects in ClinicalChemistry was formed in 1978 to study certain aspects in thisproblem. The first chairman was Prof. G. Siest, Nancy, France.

The members of the Committee on Drug Effects in Clinical Chemistrythat finalized the publications during 1988 were J. Breuer, Gelsenkirchen,DE, M.M. Galteau, Nancy, FR, P.A.G. Malya, Wilmington, Delaware, US andN. Tryding, Kristianstad, SE (Chairman). The committee was dissolvedwhen all its projects had been finalized. The following recommendationshave been published by IFCC:

Part 1.
The basic concepts.
IFCC 1984/2.
J. Clin. Chem. Clin. Biochem. 1984; 22: 271-274.
Clin. Chim. Acta 1984; 139: 215F-221F.

Part 2.
Guidelines for evaluation of analytical interference.
IFCC 1984/3.
J. Clin. Chem. Clin. Biochem. 1984; 22: 275-279.
Clin. Chim. Acta 1984; 139, 223F-230F.

Part 3.
Evaluation of biological effects of drugs.
IFCC 1988/2.
J. Clin. Chem. Clin. Biochem. 1988; 26: 169-173.
biochimica clinica 1988; 12: 972-976.
J. Biomed. Lab. Sci. 1988; 1: 35-41.

Part 4.
Clinical laboratory tests on laboratory animals during
toxicity studies.
J. Clin. Chem. Clin. Biochem. 1988; 26: 175-179.
biochimica clinica 1988; 12: 977-981.
Labmedica 1988; 5: 19-21.

Part 5.
Laboratory tests during clinical trials.
IFCC 1987/1.
J. Clin. Chem. Clin. Biochem. 1987; 25: 185-189.
biochemica clinica 1987; 11: 810-814.
Lab medica 1987-88; 4: 21-24.
Ann. Biol. Clin. 1988;46:743-769.
Analisis de laboratorio durante ensayos clinicos.
Bioquim. Clin. Latinoam. 1987; 81: 421-428.

Part 6.
Laboratory tests in monitoring drug administration.
IFCC 1988/3.
J. Clin. Chem. Clin. Biochem. 1988; 26: 181-186.
biochimica clinica 1988; 12: 983-988.
J. Biomed. Lab. Sci. 1988; 1: 51-63.

Part 7.
Data banks.
IFCC 1987/2.
J. Clin. Chem. Biochem. 1987; 25: 191-194.
biochimica clinica 1987; 11: 814-817.
Banques de données.
Inform. Scient. Biol. 1984; 10: 164-167.
Ann. Biol. Clin. 1986; 44: 70-76.
Bancos de datos efectos de drogas en quimica clinica.
Bioquim. Clin. Latinoam. 1987; 81: 429-434.

All documents have been compiled in a booklet edited by A Kallnerand N Tryding: IFCC Guidelines to the Evaluation of Drug Effectsin Clinical Chemistry.Scand. J. Clin. Lab. Invest. 1989; 49, Suppl 195: 1-28.

Interactive retrieval of clinically relevant information via Internet,data terminals, personal computers and printouts

Medical literature contains an enormous amount of references concerningdrug influences on laboratory test results. There is a real pollutionof information. Data are sometimes contradictory and the amountof data is rapidly increasing. Systematic collection and evaluationof information concerning clinically important drug interferences and effects in clinical chemistry are thereforeneeded. Information is available in several disciplines and itis necessary for people working in many different fields to learnfrom each other. A major problem is to evaluate the clinical importanceof the reports. Many reports lack significance for clinical decisionsand the reports may even be misleading. The task of establishinga reliable database is enormous and a world wide collaborationis needed. For 25 years we have evaluated all available literaturereferences with the intention of finding and assorting clinicallyrelevant information. This information is intended to be easilyavailable i.e. during clinical rounds and in other contacts betweenclinical chemists, pharmacists and clinicians.

All references in this volume have been studied and evaluatedin accordance with the clinical relevance of the drug effects.For complete information readers are referred to the original papers. The compiling of SWEDIS (SwedishDrug Information System) started in 1975 by the Medical ProductsAgency with the purpose to store information on all pharmaceuticalproducts on the Swedish market. Practical information on the effectof pharmacological doses of drugs on laboratory test results hasbeen added as an extra feature to SWEDIS.

Our aim is thus to mainly include effects which are judged clinicallyrelevant. To achieve this all references are evaluated from originalpapers. Some additional rules are applied and thus reports ofthe following types are excluded:

The relational database today contains:


An updated version of the database will be available on the WorldWide Web (Internet) during 1996 at the address http://www.pharmasoft.se/.For more information contact PharmaSoft AB, PO Box 1237, S-75142 Uppsala, Sweden, telephone +46 18 18 54 00, telefax +46 1810 92 00 or go to our above mentioned World Wide Web address.This database will include the additions and deletions found andexamined by the Swedish working committee responsible for thedatabase. The retrieval routines are easy to use and flexibleallowing the user to retrieve a great variety of information.Examples of lists that can be produced are:

The database is continuously reevaluated and updated.

Newly registered drugs are automatically included through a linkto the register of pharmaceutical specialities in SWEDIS.

For those with access to the adverse drug reaction database inSWEDIS it is possible to compare and verify published drug effectson laboratory tests with case reports of adverse drug reactions.

In many hospitals patient information (i.e. laboratory test results,diagnoses and medication) is stored in information systems, whichcan be combined with our database. This is also available in aPC version.

Sources of information

Literature references on drug effects in clinical chemistry haveexpanded enormously in the last years.

The fourth edition of the collection of references "Effectsof Drugs on Laboratory Tests" by D.S. Young was published by AACC PressUSA in 1995.

The book "Clinical Guide to Laboratory Tests" Ed NorbertW. Tietz 3rd edition, W.B. Saunders Co, Philadelphia (USA) 1995contains important information on influences from different sourcesincluding drugs on laboratory test results.

In this new book, relevant documentation is added from medicaland pharmacological books, reviews and journals. For all analyticalinterferences, brief comments regarding the type of procedurewhich is influenced are included. The total number of differententries included has increased to 18,900. Only 6% concern analyticalin vitro interferences, the rest are in vivo effects. The numberof different drugs and tests included are 950 and 370 respectively.The literature list contains 6,500 references. However, it isnot only the number of references that counts. Most importantis the quality of the information. Several hundred drug effectswith unverified documentation have been omitted in this editionof the book. Information about old tests (e.g. glucose determinationwith reduction methods) that are no longer used has also beentaken away.

We have tried to find well-documented original reports and criticalreviews. A good example is a valuable review (5193) that concernsthe effect of commonly prescribed drugs on blood lipids. Morethan 500 literature articles are reviewed and the available informationon effects needed to be considered in clinical practice is summarized.

The problems concerning the decisions if a drug has an effector not on laboratory tests in clinical practice can be illustratedby a recent publication (6492). A patient was found to have lowserum TSH values due to the intake of acetylsalicylic acid. However,if you read the literature carefully, there is no real proof ofthat this drug in practice lowers S-TSH. In fact the natural causefor the low value was the underlying disease of the patient accordingto the literature and clinical experience. In our lists we haveincluded several other "no-effects" of drugs whenthere has been a clinical suspicion but no scientific evidence.

Our problem can also be exemplified by a story from my home town.The military headquarters in Kristianstad are responsible forall soldiers in southern Sweden. A computer is used for registrationof all names and addresses. On one occasion the wrong file wasused and a mobilisation letter sent to 3000 persons who were allregistered as dead. This was of course a great mistake but itis still worth noting that 13 soldiers really did come.

Another story concerns a teacher who had a group of students whobehaved very badly. His collegues contacted him and said we regretthat you must take care of these students. However, the teachersaid that it does not matter, I take my revenge. His colleguesasked: How is that possible? The teachers answer was: I teachthem wrong things. In this 7th edition of the book we have revisedall known errors. However, you can never be safe.

We are aware of the fact that all the information we have collectedmight not be correct in all details. To improve the quality wewould be happy to receive opinions from the users of our databaseand this book.

Nils Tryding
Box 57
S-296 21 Åhus, Sweden
E-mail: nils@tryding.se
Telefax +46 44 24 12 60