BIOPHARMACEUTICS & DRUG DISPOSITION, VOL. 12, 505-514 (1991)
PHARMACOKINETICS OF AZITHROMYCIN AFTER SINGLE ORAL DOSING OF EXPERIMENTAL ANIMALS D.
DAVILA* AND LIDIJA KOLACNY-BABIC
PIiva Research Institute, Laboratory of Experimental Medicine, IL Ribara 89,41000 Zagreb, Yugoslavia
F. PLAVSIC Rebro Clinical Hospital, Department of Clinical Pharmacology, Zagreb, Yugoslavia.
ABSTRACT Azithromycin, a macrolide antibiotic with an enhanced antimicrobial spectrum, was found to have a longer half-life than erythromycin, with marked tissue penetration. The pharmacokinetics of azithromycin after oral administration were compared with those of erythromycin in rats (200 mg kg-') and rabbits (80 mg kg-I). Concentrations of azithromycin in liver, lung, kidney, ileum, and brain were higher than serum concentrations. The slow decline in tissue concentrations was evident from the biphasic elimination profile. Thus, advantageous pharmacokinetic properties and the broader antimicrobial spectrum of azithromycin relative to erythromycin appear to further support its therapeutic potential. KEY WORDS
Azithromycin Erythromycin Pharmacokinetics
Tissue levels
INTRODUCTION Azithromycin is a novel 15-membered marolide antibiotic.'**It has a broader antimicrobial spectrum and longer half-life than erythr~mycin.~-~ The present study provides further characterization of the pharmacokinetics and tissue penetration properties of this semi-synthetic macrolide.
MATERIALS AND METHODS Male Fisher rats (n = 8) weighing 160-180 g were acclimatized 7 days prior to treatment. Following administration of azithromycin or erythromycin, the animals were caged in groups of four, with free access to tap water and food (standard pellets for laboratory rats). Chinchilla female rabbits (n = 10)
* Addressee for correspondence 0142-278219 1lO70505-10$05.00 01991 by John Wiley & Sons, Ltd.
Received 7 June 1990 Revised 28 February 1991
506
D. DAVILA,
F. PLAVSIC AND L.
KOLAI~NY-BABIC
weighing 4.0-6.3 kg were housed separately in cages and fed with standard food for laboratory rabbits. Water (tap) and food were offered ad libitum. Azithromycin (Pliva) was suspended in a 0.5 per cent solution of methylcellulose and gavaged to rats ( n = 8 ) in a dose of 200 mg kg-'. Another group of animals (n = 8) received the same dose of erythromycin (erythromcyin estolate, Pliva) also by gavage. Rabbits (n = 10) received azithromycin or erythromycin in oral capsules in a dose of 80 mg kg-I. Rats were sacrificed and blood samples collected before (0 h) and at the following times after drug administration: 1, 2, 3, 6, 12, 24, and 48 h. All samples were centrifuged at 3000 X g at room temperature, and the serum was separated and stored at -20", until determination of azithromycin or erythromycin. At the same time intervals, the organs (liver, kidney, lung, ileum, and brain) were removed and prepared for determination of antibiotic content. Urinary excretion was assessed in one group ( n = 8) of rats. For this purpose, the rats already given a dose were kept individually in metabolic cages allowing collection of urine. Urine samples were collected at 6, 12, 24, and 48 h after administration of azithromycin or erythromycin. Blood from rabbits was sampled from the marginal ear vein before (0 h) and at the following times after dosing: 0.5, 1,2 ,4 ,8 , 12, and 24 h. Azithromycin and erythromycin, in sera, urines, and water tissue extracts, were determined by an agar diffusion assay using Micrococcus luteus ATCC 9341 as the test microorganism. The microbiological assay was performed using a standard The following compartmental and model-independent parameters were determined from serum concentrations according to the usual relationslo using a Hewlett-Packard 85 PC: peak concentration (C,,,), observed time to peak concentration (t,,,), time taken for the drug to reach the systemic circulation (t,,), absorption half-life (tli2abs)r distribution half-life (tli2J, elimination half-life (tli2el), volume of distribution (Vd), area under concentration-time curve (in serum or tissue) from zero to infinity calculated by the trapezoidal method (AUCo-,). RESULTS Samples of serum, urine, and tissue obtained before oral administration of azithromycin or erythromycin (0 h) showed no detectable antibiotic activity. Figure 1 shows mean concentrations of azithromycin and erythromycin found in rat serum. In Table 1, relevant pharmacokinetic parameters are presented. After oral administration, both antibiotics were observed to achieve their maximal serum concentrations after 2-3 h. The values for the elimination half-life, V, and AUC%, for azithromycin were higher than those for erythromycin. Excretion rates and related parameters are presented in Table 2. The excretion of azithromycin was greater than that of erythromycin in each of the four
507
AZITHROMYCIN
1
2
3
hours
6
12
24
48
Figure 1. Mean (n = 8) concentrations of azithromycin and erythromycin in serum of rats given a single oral dose of 200 mg kg-'
Table 1. Mean (n = 8) pharmacokinetic parameters for oral application of azithromycin and erythromycin following a dose of 200 mg kg-' to rats Pharmacokinetic parameter+(unit)
Azithromycin
Erythromycin
0.49 1.2 1.17 30.3 14.1 2.44 1.93 1 16.0 41.6 0.98
0.33 0.65 0.92 -
10.1
1.44 1.87 109.2 29.1 0.99
For definition see Material and Methods.
sampling periods, but especially in the fraction collected over the first period (0-6 h). The mean concentrations of azithromycin and erythromycin in the liver, kidney, lung, ileum, and brain of rats after oral doses of 200 mg kg-' are shown in Table 3. The concentrations of azithromycin in each case were significantly higher than those obtained similarly for erythromycin. At the same time, azithromycin concentrations in all tissues (except brain) examined were signifi-
D. DAVILA, F. PLAVSIC AND
508
L.
KOLA~NY-BABIC
Table 2. Mean (n = 8) urinary excretion of azithromycin (A) and erythromycin (E) after a single oral dose of 200 mg kg-' to rats Collection interval (h)
Amount excreted (mg) A E 512 296 177 85
0-6 612 12-24 24-48
187 181 111 57
Excretion rate (mg/min) A E 85.3 49.3 14.7 3.5
Cumulative total (mg) A E
31.2 30.2 9.3 2.4
512 808 985 I070
187 368 479 536
Table 3. Mean azithromycin (A) and erythromycin (E) concentration in rat ( n = 8 ) tissue after oral application (200 mg kg-') ~
~~~~
Tissue
~
Antibiotic 2
Concentration (mg 1-I) at time (h)* 12 24
48
Liver
A E
959 f 77 91 f 9
456 f 151 12f3
158 f 143 1.3 f0.09
63 f 30 0.1 f 0 . 1
Kidney
A E
483 f 123 69 f 37
389 f 187 30f21
281 f 176 2.8 f0.3
99 f 4 3 0.2 f0.13
Lung
A E
459 f 173 68f7
401 f 77 41 f 2 5
233 f 179 5.3 f 2.7
26f 17 0.24 f0.26
Ileum
A E
1989 f 9 3 1 74 f 16
551 f 351 27 f 24
309 f267 4.6 f 2.1
39f 15 0.05 f0.1
Brain
A E
3.2 f 1.6 0.19 f 0.2
3.8 f 1.4 0.17 f 0.2
2.9 f 1.2 0
0-3 f0.1 0
* Mean f standard deviation of eight determinations. cantly higher than those in serum. The brain erythromycin concentration after 24 h was beyond detection. Ratios of tissue to serum concentration following 200 mg kg - I doses of azithromycin and erythromycin given to rats are presented in Table 4. The values for erythromycin were much lower than those obtained for azithromycin. The relation between time (x) and ratio (y) is expressed as a polynomial with a degree of three or more 0,= a bx + C X O . ~ + dx0'25)(Table
+
5).
Values of pharmacokinetic parameters were derived from rat tissue concentrations; they are summarized in Table 6. In each case, they were considerably higher for azithromycin than for erythromycin. Table 7 shows pharrnacokinetic data derived by simulation of oral, multiple dosing of rats with azithromycin and erythromycin (2.5 mg kg-'), respectively, at 4-hourly intervals. Figure 2 shows azithromycin and erythromycin concentrations values in rabbit serum after single doses of 80 mg kg-I; the corresponding pharmacokinetic
509
AZITHROMYCIN
Table 4. Azithromycin (A) and erythromycin (E) tissue/serum ratios in rats (n = 8) after oral application of 200 mg kg-I Tissue
Antibiotic
Tissue/serum ratios at time (h) 12 24
2 Liver
48
A E
780 53
1381 28
480 5
331 2
Kidney
A E
303 37
1171 59
395 11
46 1 2
Lung
A E
374 37
1261 101
705 21
138 3
Ileum
A E
1617 40
1682 60
938 18
207 0.6
Brain
A E
2.6 0.1
11.5 0.4
8.8 0
1.5 0
Table 5. Coefficients of polynomial regression of tissue/serum concentration ratios for oral doses (200 mg kg-I) of azithromycin (A) and erythromycin (E) to rats Tissue
+ + cxO,’+ dx0,25
a
y = a bx b
C
d
An tibiotic
Liver
A E
2 185 195
546 9
13621 196
34 307 427
Kidney
A E
7 622 87
104 21
3 738 548
11 010 1381
Lung
A E
13 226 2 026
243 49
7 193 1 224
19581 1830
Ileum
A E
9 324 827
23 1 21
6 490 548
16454 1384
Brain
A
76
33
105
0.69
parameters are presented in Table 8. Azithromycin concentrations in rabbit sera were higher than those of erythromycin. Maximal azithromycin concentration was achieved at about 60 min after dosing, whereas for erythromycin it took about 120 min. The AUCs and volumes of distribution are also notably different. DISCUSSION In this study, performed with two animal species, azithromycin demonstrated a markedly long half-life and extensive tissue penetration. After oral adminis-
510
PLAVSIC AND
D. DAVILA, F.
L. KOLACNY-BABIC
Table 6. Mean (n = 8) pharmacokinetic parameters in various tissues of the rat after oral application of azithromycin (A) and erythromycin (E) (200 mg kg-I) Tissue
Antibiotic
Liver Kidney Lung Ileum Brain
Pharmacokinetic parametert (unit) t %el
t'habs
G a x
(h)
(h)
(mg 1-I)
AUCkinf (mg*h 1-I)
A E
13.1
962 5.2
962 92
17000 579
A E
18
-
538 69
23 268 673
752 95
13 173 960
1992 77
23 429 718
-
5.2
-
A E
8.9
-
-
4.9
A E
9.2
-
-
4.3
A
7.2
-
0.5
131
For definition see Material and Methods.
Table 7. Mean (n = 8) steady-state concentrations of azithromycin and erythromycin in the rat after oral doses of 2.5 mg kg-' at 4-hourly intervals Tissue
cmx
Time to steady state (days)
Clearance (h)
0.22 40 35 27 60 0.16
10 3 5 2 3 2
300 91 125 62 65 50
3 1 1
93 38 36 35 31
Steady-state concentration
Girl
6% 1 - 9
Azithromycin Serum Liver Kidney Lung Ileum Brain
0.2 27 30 24 40 0.13
Serum Liver Kidney Lung Ileum
0.04
Erythromycin
0.82 1 1.75 1.4
0.055 1.75 1.5 2.2 1.8
1
1
tration of azithromycin to rats, the ratio of tissue to serum concentrations (Table 4) was, in each case, much higher than for erythromycin. Accordingly, it may be concluded that azithromycin is stored to a greater extent in the tissues examined than is erythromycin. The regression of concentration ratios for azithromycin and erythromycin after oral administration (Table 5) to rats appears to follow a biexponential
51 1
AZITHROMYCIN 0.7
0.5
0.3
0.1
0.5
I
hours
8
12
tl
Figure 2. Mean (n = 10) concentrations of azithromycin and erythromycin in serum of rabbits give a single oral dose of 80 mg kg-'
Table 8. Mean (n = 10) pharmacokinetic parameters for azithromycin and erythromycin after administration of doses of 80 mg kg-' to rabbits. Pharmacokinetic parameterst (unit) flag (h)
(h) t,, (h) t'he.1 (h) tmax (h) Cmax (mg I-') A (mg 1 - I ) B (mg 1-I) AUC%id(mg*h 1-I) VdS0 kg-9 t'habs
Azithromycin
Erythromycin
0 0.26 0.19 29-1 0.72 0.69 125 0.14 20 57 1
0.21 0.71 0.85 12.8 1.8 0.12 0.82 0.105 4 762
For definition see Material and Methods.
* Determined assuming absolute bioavailability of approximately
one.
curve. However, as there was no reason to believe that the biexponential equation applied, the data were analysed as a polynomial of the third degree and, in fact, this produced the best-fitting curve. Such a form would be expected
512
D.
DAVILA, F. PLAVSIC AND L. K O L A ~ N Y - B A B I ~
tissuekerum ratio 1800 1600
1
D-------\
1400 1200 1000 800 600 400 200
'A
0 '
J
2
12
hours
24
48
Figure 3. Presentation of shapes of the concentration tissue/serum ratio versus time curves for ileum, liver, lung, and kidney in rats (n = 8) after oral a plication of azithromycin following a dose of 200 mg kg-
P
if azithromycin elimination from serum is determined mainly by its release from depot tissue and not on excretion by the kidney, which is a rather fast process. At the same time it can be seen that after oral administration of the drugs, the concentration tissue/serum ratio versus time profiles were similar for all tissues (Figure 3). Pharmacokinetic parameters given in Table 1 show that azithromycin is very quickly distributed from serum to tissue (or organs) and then released very slowly from this depot to serum. Deposition is less prominent with erythromycin; hence, a two-compartment model was used for pharmacokinetic calculations. The pharmacokinetic parameters (Table 6 ) indicate marked deposition of azithromycin in the various organs; in other words,.the drug is delivered quickly to the depot but released very slowly. For erythromycin, such an effect was not evident; in fact, release from the depot was faster than elimination of the drug from the body. Thus deposition influences only the volume of distribution in the body. Such results are consistent with the understanding that antibiotic efficacy (antibacterial activity) in vivo is associated directly with concentration achieved in tissues.'*J2In our investigation, we did not differentiate between free and bound azithromycin in tissues. We believe this to be justified because azithromycin enters the tissues in the free (active) form although it is bound rapidly to sites in cell membranes and subcellular structures. The total concen-
AZITHROMYCIN
513
tration in tissues, i.e. free and bound form, will thus reflect the concentration of the active drug. Thus the activity of azithromycin can be correlated with its tissue c o n c e n t r a t i ~ n . ~ ~ ~ ~ In order to further examine the accumulation of the drugs in the depots, azithromycin and erythromycin concentrations in serum and various tissues were determined after multiple dosing; the regimen was 2.5 mg kg-I dose at 4-hourly intervals. Such a regimen corresponds more or less to that for erythromycin when used to treat humans (Table 7). Steady-state azithromycin concentrations in kidneys were much higher (160 times) than the maximal steady-state concentration in serum. This is attributable to the slow azithromycin renal clearance (125 h) after oral administration to steady state. It is somewhat less pronounced for other tissues, but, in some cases, deposition was significant. Erythromycin did not show a tendency to accumulate; C,, values were low and clearance rapid. Our results showed that azithromycin penetrates tissue well and is then eliminated relatively slowly. Thus it achieves and maintains high levels in the organs. Most infections are localized in tissues or organs; the concentration of antibiotic in blood or serum is not then the most appropriate parameter for assessing the activity of an antibiotic. Tissue concentrations are of greater clinical relevance. Based on its pharmacokinetics, the regime for the clinical use of azithromycin, is often rather very short; for some infections, a single dose is sufficient (unpublished results). Azithromycin concentrations after single oral administration to rabbits were higher than those obtained similarly for erythromycin (Figure 2, Table 8). Notable were differences in the values obtained for AUCs and volumes of distribution. Our preliminary results obtained in dogs showed that volumes of distribution of azithromycin were significantly lower than in rodents but maximal concentrations were much higher. Thus, our results are in agreement with data obtained by Girard et aL6 and Shepard and FalkneP in mice, dogs, and monkeys. In summary, azithromycin, a new semi-syntheticmacrolide antibiotic, quickly reaches the general circulation after oral administration. Maximal serum concentrations were reached in 0.5-3h. Its serum half-life was about 15 h. Azithromycin quickly attains high concentrations in the various organs examined, but is cleared rather slowly. Thus, the results obtained have at least two important and practical implications. The evident accumulation means that azithromycin concentrations in organs are relatively high. Accordingly, since the observed concentrations were greater than minimal inhibitory concentrations for various microorganisms, azithromycin can act with greater effect than antibiotics such as erythromycin, which are eliminated relatively quickly. Thus, accumulation of azithromycin in organs may prove useful in the therapy of various infections. Furthermore, this property of azithromycin points to the possibility of decreasing the level and frequency of doses. This is particularly promising because, up to now,
514
D. DAVILA,
F.
PLAVSIC
AND
L. KOLA~NY-BABIC
neither the accumulation of azithromycin nor its relatively long residence there have been observed to induce any untoward effects. When the drug was administered for several months (toxicity tests in rats, rabbits, and dogs) in amounts greatly exceeding the therapeutic dose, no damaging effects on the animals were observed. Accordingly, with significantly lower therapeutic doses, side-effects are still less likely to occur.
REFERENCES 1. S. DjokiC, G. Kobrehel, G. Lazarevski, N. Lopotar, Z. TamburaSev, B. Kamenar, A. Nag1 and I. Vickovid, J. Chem. SOC.Perkin Transact., I, 1881 (1986). 2. S. DjokiC, G. Kobrehel, N. Lopotar, B. Kamenar, A. Nag1 and D. MrvoS, J. Chem. Res., (M), 1239 (1988). 3. T. TambiC, S. Djokid, D. Davila and L. KolaEny-BabiC,Acta Pharm. Jugosl., 39,233 (1989). 4. J. Retsema, A. Girard, W. Schekly, M. Manousos, M. Anderson, G. Bright, R. Borovoy, L. Brennan and R. Mason, Antimicrob. Agents Chemother.,31,1939 (1987).
5 . D. Davila, F. PlavSiC, Z. Mubrin, L. KolaEny-BabiC and M. Gokv, Proceedings of the 5th Czechoslovak Congress of Infectious and Parasitic Diseases. 0. Balint and I. BakoS (Eds), Bratislava, 1989, p. 1 1 1. 6. A. E. Girard, D. Girard, A. R. English, T. D. Gootz, C. R. Cimochowski, J. A. Faiella, S. L. Haskell and J. A. Retsema, Antimicrob. Agents Chemother., 31, 1948 (1987). 7. D. Davila and F. PlavSiC, Pharmas 89: International Symposium of Pharmacokinetic Modeling and Simulation, Portoroi, 1989, p. 35. 8. S. M. Finegold, J. W. Martin, B. Bailey and S. Scott (Eds), Diagnostic Microbiology, Mosby Co., St. Louis-Toronto-London, 1982, p. 554. 9. L. W. Hewit and M. C. McHenry, Med. Clin. North Amer., 62, 11 19 (1978). 10. W. A. Ritschel, Handbook ofBasic Pharmacokinetics,Drug Intelligence Publications, Hamilton, 1982. 11. M. G. Bergeron, Clin. Biochem., 19,90 (1986). 12. E. Goormans, A. Dalhoff, B. Kazzas and J. Branolte, Chemotherapy, 32,7 (1986). 13. K. H. Chan, J. D. Swarts, W. J. Doyle, K. Taupowpong and D. R. Kardatzke, Arch. Otolaryngol. HeadNeck Surg., 114, 1266 (1988). 14. A. E. Girard, D. Girard and J. A. Retsema, J. Antimicrob. Chemother., 25, suppl. A, 61 (1990). 15. R. C. Johnson, C. Kodner, M. Russell and D. Girard, J. Antimicrob. Chemother., 25, suppl. A, 33 (1990). 16. R. M. Shepard and F. C. Falkner, J. Antimicrob. Chemother., 25, suppl. A, 49 (1990).
PHARMACOKINETICS OF AZITHROMYCIN AFTER SINGLE ORAL DOSING OF EXPERIMENTAL ANIMALS D.
DAVILA* AND LIDIJA KOLACNY-BABIC
PIiva Research Institute, Laboratory of Experimental Medicine, IL Ribara 89,41000 Zagreb, Yugoslavia
F. PLAVSIC Rebro Clinical Hospital, Department of Clinical Pharmacology, Zagreb, Yugoslavia.
ABSTRACT Azithromycin, a macrolide antibiotic with an enhanced antimicrobial spectrum, was found to have a longer half-life than erythromycin, with marked tissue penetration. The pharmacokinetics of azithromycin after oral administration were compared with those of erythromycin in rats (200 mg kg-') and rabbits (80 mg kg-I). Concentrations of azithromycin in liver, lung, kidney, ileum, and brain were higher than serum concentrations. The slow decline in tissue concentrations was evident from the biphasic elimination profile. Thus, advantageous pharmacokinetic properties and the broader antimicrobial spectrum of azithromycin relative to erythromycin appear to further support its therapeutic potential. KEY WORDS
Azithromycin Erythromycin Pharmacokinetics
Tissue levels
INTRODUCTION Azithromycin is a novel 15-membered marolide antibiotic.'**It has a broader antimicrobial spectrum and longer half-life than erythr~mycin.~-~ The present study provides further characterization of the pharmacokinetics and tissue penetration properties of this semi-synthetic macrolide.
MATERIALS AND METHODS Male Fisher rats (n = 8) weighing 160-180 g were acclimatized 7 days prior to treatment. Following administration of azithromycin or erythromycin, the animals were caged in groups of four, with free access to tap water and food (standard pellets for laboratory rats). Chinchilla female rabbits (n = 10)
* Addressee for correspondence 0142-278219 1lO70505-10$05.00 01991 by John Wiley & Sons, Ltd.
Received 7 June 1990 Revised 28 February 1991
506
D. DAVILA,
F. PLAVSIC AND L.
KOLAI~NY-BABIC
weighing 4.0-6.3 kg were housed separately in cages and fed with standard food for laboratory rabbits. Water (tap) and food were offered ad libitum. Azithromycin (Pliva) was suspended in a 0.5 per cent solution of methylcellulose and gavaged to rats ( n = 8 ) in a dose of 200 mg kg-'. Another group of animals (n = 8) received the same dose of erythromycin (erythromcyin estolate, Pliva) also by gavage. Rabbits (n = 10) received azithromycin or erythromycin in oral capsules in a dose of 80 mg kg-I. Rats were sacrificed and blood samples collected before (0 h) and at the following times after drug administration: 1, 2, 3, 6, 12, 24, and 48 h. All samples were centrifuged at 3000 X g at room temperature, and the serum was separated and stored at -20", until determination of azithromycin or erythromycin. At the same time intervals, the organs (liver, kidney, lung, ileum, and brain) were removed and prepared for determination of antibiotic content. Urinary excretion was assessed in one group ( n = 8) of rats. For this purpose, the rats already given a dose were kept individually in metabolic cages allowing collection of urine. Urine samples were collected at 6, 12, 24, and 48 h after administration of azithromycin or erythromycin. Blood from rabbits was sampled from the marginal ear vein before (0 h) and at the following times after dosing: 0.5, 1,2 ,4 ,8 , 12, and 24 h. Azithromycin and erythromycin, in sera, urines, and water tissue extracts, were determined by an agar diffusion assay using Micrococcus luteus ATCC 9341 as the test microorganism. The microbiological assay was performed using a standard The following compartmental and model-independent parameters were determined from serum concentrations according to the usual relationslo using a Hewlett-Packard 85 PC: peak concentration (C,,,), observed time to peak concentration (t,,,), time taken for the drug to reach the systemic circulation (t,,), absorption half-life (tli2abs)r distribution half-life (tli2J, elimination half-life (tli2el), volume of distribution (Vd), area under concentration-time curve (in serum or tissue) from zero to infinity calculated by the trapezoidal method (AUCo-,). RESULTS Samples of serum, urine, and tissue obtained before oral administration of azithromycin or erythromycin (0 h) showed no detectable antibiotic activity. Figure 1 shows mean concentrations of azithromycin and erythromycin found in rat serum. In Table 1, relevant pharmacokinetic parameters are presented. After oral administration, both antibiotics were observed to achieve their maximal serum concentrations after 2-3 h. The values for the elimination half-life, V, and AUC%, for azithromycin were higher than those for erythromycin. Excretion rates and related parameters are presented in Table 2. The excretion of azithromycin was greater than that of erythromycin in each of the four
507
AZITHROMYCIN
1
2
3
hours
6
12
24
48
Figure 1. Mean (n = 8) concentrations of azithromycin and erythromycin in serum of rats given a single oral dose of 200 mg kg-'
Table 1. Mean (n = 8) pharmacokinetic parameters for oral application of azithromycin and erythromycin following a dose of 200 mg kg-' to rats Pharmacokinetic parameter+(unit)
Azithromycin
Erythromycin
0.49 1.2 1.17 30.3 14.1 2.44 1.93 1 16.0 41.6 0.98
0.33 0.65 0.92 -
10.1
1.44 1.87 109.2 29.1 0.99
For definition see Material and Methods.
sampling periods, but especially in the fraction collected over the first period (0-6 h). The mean concentrations of azithromycin and erythromycin in the liver, kidney, lung, ileum, and brain of rats after oral doses of 200 mg kg-' are shown in Table 3. The concentrations of azithromycin in each case were significantly higher than those obtained similarly for erythromycin. At the same time, azithromycin concentrations in all tissues (except brain) examined were signifi-
D. DAVILA, F. PLAVSIC AND
508
L.
KOLA~NY-BABIC
Table 2. Mean (n = 8) urinary excretion of azithromycin (A) and erythromycin (E) after a single oral dose of 200 mg kg-' to rats Collection interval (h)
Amount excreted (mg) A E 512 296 177 85
0-6 612 12-24 24-48
187 181 111 57
Excretion rate (mg/min) A E 85.3 49.3 14.7 3.5
Cumulative total (mg) A E
31.2 30.2 9.3 2.4
512 808 985 I070
187 368 479 536
Table 3. Mean azithromycin (A) and erythromycin (E) concentration in rat ( n = 8 ) tissue after oral application (200 mg kg-') ~
~~~~
Tissue
~
Antibiotic 2
Concentration (mg 1-I) at time (h)* 12 24
48
Liver
A E
959 f 77 91 f 9
456 f 151 12f3
158 f 143 1.3 f0.09
63 f 30 0.1 f 0 . 1
Kidney
A E
483 f 123 69 f 37
389 f 187 30f21
281 f 176 2.8 f0.3
99 f 4 3 0.2 f0.13
Lung
A E
459 f 173 68f7
401 f 77 41 f 2 5
233 f 179 5.3 f 2.7
26f 17 0.24 f0.26
Ileum
A E
1989 f 9 3 1 74 f 16
551 f 351 27 f 24
309 f267 4.6 f 2.1
39f 15 0.05 f0.1
Brain
A E
3.2 f 1.6 0.19 f 0.2
3.8 f 1.4 0.17 f 0.2
2.9 f 1.2 0
0-3 f0.1 0
* Mean f standard deviation of eight determinations. cantly higher than those in serum. The brain erythromycin concentration after 24 h was beyond detection. Ratios of tissue to serum concentration following 200 mg kg - I doses of azithromycin and erythromycin given to rats are presented in Table 4. The values for erythromycin were much lower than those obtained for azithromycin. The relation between time (x) and ratio (y) is expressed as a polynomial with a degree of three or more 0,= a bx + C X O . ~ + dx0'25)(Table
+
5).
Values of pharmacokinetic parameters were derived from rat tissue concentrations; they are summarized in Table 6. In each case, they were considerably higher for azithromycin than for erythromycin. Table 7 shows pharrnacokinetic data derived by simulation of oral, multiple dosing of rats with azithromycin and erythromycin (2.5 mg kg-'), respectively, at 4-hourly intervals. Figure 2 shows azithromycin and erythromycin concentrations values in rabbit serum after single doses of 80 mg kg-I; the corresponding pharmacokinetic
509
AZITHROMYCIN
Table 4. Azithromycin (A) and erythromycin (E) tissue/serum ratios in rats (n = 8) after oral application of 200 mg kg-I Tissue
Antibiotic
Tissue/serum ratios at time (h) 12 24
2 Liver
48
A E
780 53
1381 28
480 5
331 2
Kidney
A E
303 37
1171 59
395 11
46 1 2
Lung
A E
374 37
1261 101
705 21
138 3
Ileum
A E
1617 40
1682 60
938 18
207 0.6
Brain
A E
2.6 0.1
11.5 0.4
8.8 0
1.5 0
Table 5. Coefficients of polynomial regression of tissue/serum concentration ratios for oral doses (200 mg kg-I) of azithromycin (A) and erythromycin (E) to rats Tissue
+ + cxO,’+ dx0,25
a
y = a bx b
C
d
An tibiotic
Liver
A E
2 185 195
546 9
13621 196
34 307 427
Kidney
A E
7 622 87
104 21
3 738 548
11 010 1381
Lung
A E
13 226 2 026
243 49
7 193 1 224
19581 1830
Ileum
A E
9 324 827
23 1 21
6 490 548
16454 1384
Brain
A
76
33
105
0.69
parameters are presented in Table 8. Azithromycin concentrations in rabbit sera were higher than those of erythromycin. Maximal azithromycin concentration was achieved at about 60 min after dosing, whereas for erythromycin it took about 120 min. The AUCs and volumes of distribution are also notably different. DISCUSSION In this study, performed with two animal species, azithromycin demonstrated a markedly long half-life and extensive tissue penetration. After oral adminis-
510
PLAVSIC AND
D. DAVILA, F.
L. KOLACNY-BABIC
Table 6. Mean (n = 8) pharmacokinetic parameters in various tissues of the rat after oral application of azithromycin (A) and erythromycin (E) (200 mg kg-I) Tissue
Antibiotic
Liver Kidney Lung Ileum Brain
Pharmacokinetic parametert (unit) t %el
t'habs
G a x
(h)
(h)
(mg 1-I)
AUCkinf (mg*h 1-I)
A E
13.1
962 5.2
962 92
17000 579
A E
18
-
538 69
23 268 673
752 95
13 173 960
1992 77
23 429 718
-
5.2
-
A E
8.9
-
-
4.9
A E
9.2
-
-
4.3
A
7.2
-
0.5
131
For definition see Material and Methods.
Table 7. Mean (n = 8) steady-state concentrations of azithromycin and erythromycin in the rat after oral doses of 2.5 mg kg-' at 4-hourly intervals Tissue
cmx
Time to steady state (days)
Clearance (h)
0.22 40 35 27 60 0.16
10 3 5 2 3 2
300 91 125 62 65 50
3 1 1
93 38 36 35 31
Steady-state concentration
Girl
6% 1 - 9
Azithromycin Serum Liver Kidney Lung Ileum Brain
0.2 27 30 24 40 0.13
Serum Liver Kidney Lung Ileum
0.04
Erythromycin
0.82 1 1.75 1.4
0.055 1.75 1.5 2.2 1.8
1
1
tration of azithromycin to rats, the ratio of tissue to serum concentrations (Table 4) was, in each case, much higher than for erythromycin. Accordingly, it may be concluded that azithromycin is stored to a greater extent in the tissues examined than is erythromycin. The regression of concentration ratios for azithromycin and erythromycin after oral administration (Table 5) to rats appears to follow a biexponential
51 1
AZITHROMYCIN 0.7
0.5
0.3
0.1
0.5
I
hours
8
12
tl
Figure 2. Mean (n = 10) concentrations of azithromycin and erythromycin in serum of rabbits give a single oral dose of 80 mg kg-'
Table 8. Mean (n = 10) pharmacokinetic parameters for azithromycin and erythromycin after administration of doses of 80 mg kg-' to rabbits. Pharmacokinetic parameterst (unit) flag (h)
(h) t,, (h) t'he.1 (h) tmax (h) Cmax (mg I-') A (mg 1 - I ) B (mg 1-I) AUC%id(mg*h 1-I) VdS0 kg-9 t'habs
Azithromycin
Erythromycin
0 0.26 0.19 29-1 0.72 0.69 125 0.14 20 57 1
0.21 0.71 0.85 12.8 1.8 0.12 0.82 0.105 4 762
For definition see Material and Methods.
* Determined assuming absolute bioavailability of approximately
one.
curve. However, as there was no reason to believe that the biexponential equation applied, the data were analysed as a polynomial of the third degree and, in fact, this produced the best-fitting curve. Such a form would be expected
512
D.
DAVILA, F. PLAVSIC AND L. K O L A ~ N Y - B A B I ~
tissuekerum ratio 1800 1600
1
D-------\
1400 1200 1000 800 600 400 200
'A
0 '
J
2
12
hours
24
48
Figure 3. Presentation of shapes of the concentration tissue/serum ratio versus time curves for ileum, liver, lung, and kidney in rats (n = 8) after oral a plication of azithromycin following a dose of 200 mg kg-
P
if azithromycin elimination from serum is determined mainly by its release from depot tissue and not on excretion by the kidney, which is a rather fast process. At the same time it can be seen that after oral administration of the drugs, the concentration tissue/serum ratio versus time profiles were similar for all tissues (Figure 3). Pharmacokinetic parameters given in Table 1 show that azithromycin is very quickly distributed from serum to tissue (or organs) and then released very slowly from this depot to serum. Deposition is less prominent with erythromycin; hence, a two-compartment model was used for pharmacokinetic calculations. The pharmacokinetic parameters (Table 6 ) indicate marked deposition of azithromycin in the various organs; in other words,.the drug is delivered quickly to the depot but released very slowly. For erythromycin, such an effect was not evident; in fact, release from the depot was faster than elimination of the drug from the body. Thus deposition influences only the volume of distribution in the body. Such results are consistent with the understanding that antibiotic efficacy (antibacterial activity) in vivo is associated directly with concentration achieved in tissues.'*J2In our investigation, we did not differentiate between free and bound azithromycin in tissues. We believe this to be justified because azithromycin enters the tissues in the free (active) form although it is bound rapidly to sites in cell membranes and subcellular structures. The total concen-
AZITHROMYCIN
513
tration in tissues, i.e. free and bound form, will thus reflect the concentration of the active drug. Thus the activity of azithromycin can be correlated with its tissue c o n c e n t r a t i ~ n . ~ ~ ~ ~ In order to further examine the accumulation of the drugs in the depots, azithromycin and erythromycin concentrations in serum and various tissues were determined after multiple dosing; the regimen was 2.5 mg kg-I dose at 4-hourly intervals. Such a regimen corresponds more or less to that for erythromycin when used to treat humans (Table 7). Steady-state azithromycin concentrations in kidneys were much higher (160 times) than the maximal steady-state concentration in serum. This is attributable to the slow azithromycin renal clearance (125 h) after oral administration to steady state. It is somewhat less pronounced for other tissues, but, in some cases, deposition was significant. Erythromycin did not show a tendency to accumulate; C,, values were low and clearance rapid. Our results showed that azithromycin penetrates tissue well and is then eliminated relatively slowly. Thus it achieves and maintains high levels in the organs. Most infections are localized in tissues or organs; the concentration of antibiotic in blood or serum is not then the most appropriate parameter for assessing the activity of an antibiotic. Tissue concentrations are of greater clinical relevance. Based on its pharmacokinetics, the regime for the clinical use of azithromycin, is often rather very short; for some infections, a single dose is sufficient (unpublished results). Azithromycin concentrations after single oral administration to rabbits were higher than those obtained similarly for erythromycin (Figure 2, Table 8). Notable were differences in the values obtained for AUCs and volumes of distribution. Our preliminary results obtained in dogs showed that volumes of distribution of azithromycin were significantly lower than in rodents but maximal concentrations were much higher. Thus, our results are in agreement with data obtained by Girard et aL6 and Shepard and FalkneP in mice, dogs, and monkeys. In summary, azithromycin, a new semi-syntheticmacrolide antibiotic, quickly reaches the general circulation after oral administration. Maximal serum concentrations were reached in 0.5-3h. Its serum half-life was about 15 h. Azithromycin quickly attains high concentrations in the various organs examined, but is cleared rather slowly. Thus, the results obtained have at least two important and practical implications. The evident accumulation means that azithromycin concentrations in organs are relatively high. Accordingly, since the observed concentrations were greater than minimal inhibitory concentrations for various microorganisms, azithromycin can act with greater effect than antibiotics such as erythromycin, which are eliminated relatively quickly. Thus, accumulation of azithromycin in organs may prove useful in the therapy of various infections. Furthermore, this property of azithromycin points to the possibility of decreasing the level and frequency of doses. This is particularly promising because, up to now,
514
D. DAVILA,
F.
PLAVSIC
AND
L. KOLA~NY-BABIC
neither the accumulation of azithromycin nor its relatively long residence there have been observed to induce any untoward effects. When the drug was administered for several months (toxicity tests in rats, rabbits, and dogs) in amounts greatly exceeding the therapeutic dose, no damaging effects on the animals were observed. Accordingly, with significantly lower therapeutic doses, side-effects are still less likely to occur.
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5 . D. Davila, F. PlavSiC, Z. Mubrin, L. KolaEny-BabiC and M. Gokv, Proceedings of the 5th Czechoslovak Congress of Infectious and Parasitic Diseases. 0. Balint and I. BakoS (Eds), Bratislava, 1989, p. 1 1 1. 6. A. E. Girard, D. Girard, A. R. English, T. D. Gootz, C. R. Cimochowski, J. A. Faiella, S. L. Haskell and J. A. Retsema, Antimicrob. Agents Chemother., 31, 1948 (1987). 7. D. Davila and F. PlavSiC, Pharmas 89: International Symposium of Pharmacokinetic Modeling and Simulation, Portoroi, 1989, p. 35. 8. S. M. Finegold, J. W. Martin, B. Bailey and S. Scott (Eds), Diagnostic Microbiology, Mosby Co., St. Louis-Toronto-London, 1982, p. 554. 9. L. W. Hewit and M. C. McHenry, Med. Clin. North Amer., 62, 11 19 (1978). 10. W. A. Ritschel, Handbook ofBasic Pharmacokinetics,Drug Intelligence Publications, Hamilton, 1982. 11. M. G. Bergeron, Clin. Biochem., 19,90 (1986). 12. E. Goormans, A. Dalhoff, B. Kazzas and J. Branolte, Chemotherapy, 32,7 (1986). 13. K. H. Chan, J. D. Swarts, W. J. Doyle, K. Taupowpong and D. R. Kardatzke, Arch. Otolaryngol. HeadNeck Surg., 114, 1266 (1988). 14. A. E. Girard, D. Girard and J. A. Retsema, J. Antimicrob. Chemother., 25, suppl. A, 61 (1990). 15. R. C. Johnson, C. Kodner, M. Russell and D. Girard, J. Antimicrob. Chemother., 25, suppl. A, 33 (1990). 16. R. M. Shepard and F. C. Falkner, J. Antimicrob. Chemother., 25, suppl. A, 49 (1990).
