Studies on digitalis

x.

Digitoxin metabolites in human myocardium and relationship

between myocardial and serum concentrations of digitoxin in patients on maintenance treatment The levels of digitoxin and cardioaetive metabolites were measured in 42 afrial biopsies with a R6Rb method mod(fiedfor analysis of myoeardial sampies. The mean value was 9/.0 ng Igm wet weight (SD 54.4). Myoeardial and serum eoneentrations were compared

in 23 patients; there was no signijieant correlation. The ratio o( total drug eoneentration in myoeardium and serum rangedfrom / to 38 with a mean value of5.4. Caleulatedfrom the free drug eoncentrations, the mean myoeardial serum ratio was 200, wh ich rejieets the high affinity Q( digitoxin and cardioactive metabolites to the myoeardium. The metabolie pattern ()( cardioactive and inactive metabolites (eonjugates with glucuronic and su((uric acid) was studied in autopsy sampies from left ventriel/lar myoeardium from 7 patients. Significant differences between the myocardial and serum patterns

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eardioaetive and inactive metabolites were demonstrated. The myocardium contained less unchanged digitoxin (25.7%) and more hydrolyzed (55.4%) and conjugated (54./%) metabolites than serum (57.6%, 3/.0%, and 33./ %, respeetively). Hydroxylated metabolites in myocardium (/5.S9c) were not signijicantly changed compared to serum (/0.0%).

Liv Storstein, M.D. Oslo, Norway Medical Department B, University Clinic, Rikshospitalet

Serum eoncentrations of digitalis glyeosides are currently used in the management of patients on maintenance treatment. It is therefore important to know whether serum levels mirror myocardial drug eoneentrations. lt is not possible to measure drug eoneentration at the receptor site, but determinations of total myocardial Supported by grants from the Norwegian Council on Cardiovascular Diseases, The University of Oslo, and Astra Ltd. Received for publication March 20, 1976. Accepted for publication July 15, 1976. Reprint requests to: Liv Storstein, M.D., Medical Department B, University Clinic, Rikshospitalet, Pilestredet 32, Oslo, Norway.

concentrations of digitoxin and ealculation of myocardial/serum ratios may be performed. The question is eomplieated with regard to digitoxin, which has a number of aetive and inaetive metabolites in serum. 37 The eardioaetive metabolites are protein-bound to varying degrees. 24 , 35 They differ in their affinity to the myoeardium,26 and the myoeardial pattern of metabolites may differ from that in serum. The aim of the present investigation was to determine myoeardial/serum ratios of digitoxin and eardioaetive metabolites. The metabolie patterns of both eardioaetive and inaetive, eonju395

396

Storstein

Clinical Pharmacology and Therapeutics

Table I. Accuracy of digitoxin concentration measurements in replicate sampies from human myocardium Concentration range (ng/ml)

Digitoxin added to tissue, mean ± SD (n)

Digitoxin extracted from tissue, mean ± SD (n)

0-5

4.85 ± 1.60 (10) 13.25 ± 2.98 (10) 17.35 ± 3.49 (10) 20.00 ± 3.25 (10)

3.55 ± 1.92 (20) 11.15 ± 3.53 (20) 19.75 ± 4.07 (8) 21.75 ± 3.84 (6)

10-15 15-20 >20

gated metabolites in myoeardial autopsy sampIes from patients on maintenanee treatment with digitoxin were also investigated. Material and methods

The following abbreviations for glycosides will be used: DT-3, digitoxin; DT-2, digitoxigenin-bis-digitoxoside; DT -1, digitoxigeninmono-digitoxoside; DT-O, digitoxigenin; DG3, digoxin; DG-2, digoxigenin-bis-digitoxoside; DG-I, digoxigenin-mono-digitoxoside, and DG-O, digoxigenin. Biopsy study. Biopsies from the left aurieular appendages were obtained in 42 patients during cardiopulmonary bypass and were stored at -20° C until analysis was performed. The patients were on maintenanee treatment with digitoxin (mean dose, 0.08 mg/day) and had normal serum albumin (mean value, 3.8 gm/dl) and potassium (mean value, 4.4 mEq/L). In 23 of the patients serum sampies were obtained the evening before operation. Due to the rapid fall in serum digitoxin during eardiopulmonary bypass,28 serum sampies taken simultaneously with the biopsies would show falsely low levels of digitoxin and eardioactive metabolites. Patients received their oral doses of digitoxin at 5.30 A.M. on the day of operation. Sinee drug absorption is terminated after 4 to 6 hr, myoeardial digitoxin levels should not be influeneed by absorption at the time of the biopsies. Autopsy study. Autopsy sampies were obtained from the left ventrieular myoeardium in the apical region of 7 patients. The biopsies

were too small for determination of aetive and inactive metabolites of digitoxin. Five patients died from severe heart disease, 2 males and 3 females with a mean age of 63.4 yr (range, 50 to 60 yr). One had coronary heart disease, 2 primary myoeardial disease, 1 eardiomyopathy due to amyloidosis, and 1 valvular heart disease. Laboratory values obtained shortly before death showed slightly impaired renal (mean serum creatinine 1.3 mg/dl (SD ± 0.4), urea, 71 mg/dl (±22), and hepatie (total bilirubin, 1.8 mg/ dl (± 1.0) funetions. Mean maintenanee dose of digitoxin was 0.08 mg/day (±0.02) with a me an serum level of 9.0 ng/ ml. Two patients had renal disease. One 24 yrold-woman died from a fulminant nephrotie syndrome due to lupus erythematosus. 36 She had severe hypoalbuminemia (1.1 gm per 100 ml), serum ereatinine, 2.0 mg/dl, and urea, 134 mg/dl. Her serum digitoxin level was 4.5 ng/ml on a dose of 0.1 mg/ day. The last patient was a 51-yr-old man who had chronie glomerulonephritis which was being treated with hemodialysis, and eoronary heart disease. His serum ereatinine was 9.2 mg/dl and urea, 126 mg/dl. The serum digitoxin level was 4.5 ng/mI on a digitoxin dose of 0.07 mg/day. Laboratory methods. 86Rb method. (1) The modified 86Rb method deseribed previousIy15 was used for determination of digitoxin and cardioaetive metabolites in serum. (2) The method was modified for determination of digitoxin and eardioaetive metabolites in myoeardial samples. 33 Myoeardial sampIes were homogenized in physiologie saline (+4° C) with an Ultra-Turrax homogenizer (15,000 rpm) until the suspension was very fine. Replieate sampies eontaining 50 to 100 mg myoeardium/milliliter were then taken from the homogenate with I-mI pipettes. The sampies were extraeted twice with 3 ml dichloromethane, and each time 2 ml of the extraet were transferred to a glass tube before evaporation in a water bath at 50° C. The further procedure was as for serum,15 but the standard eurve was made with a suspension of ventricular myoeardium beeause inhibition of the 8 6Rb uptake is higher with serum than with myoeardium. Two extraetions with diehloromethane proved suffieient as no digitoxin was reeovered in a third extraetion. Re-

Volume 21 Number 4

Digitoxin metabolites in human myocardium

META BOll C PAT TERN IN MYOCAROIUM

80 ACTIVE

70

INACTIVE

n -5

n- 5 60

50 40 30 20

i

I 1

10

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~

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o

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METABOLIC PATTERN IN SERUM

STEAOY STATE GROUP

80 70

ACTIVE

INACTIVE n _4

n. 9 60

50 40 30

0

or 1

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Ol,

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Studies on digitalis. X. Digitoxin metabolites in human myocardium and relationship between myocardial and serum concentrations of digitoxin in patients on maintenance treatment.

Studies on digitalis x. Digitoxin metabolites in human myocardium and relationship between myocardial and serum concentrations of digitoxin in pati...
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