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Metabolic and quantitative studies in connection with intraamniotic administration of prostaglandin F2α for induction of therapeutic abortion
Printed
iâareV a tBruinart I'
PP . 219-229, 1973
Pergamon Press
METABOLIC AND QUANTITATIVE STUDIES IN CONNECTION WITH INTRAAMNIOTIC ADMINISTRATION OF PROSTAGLANDIN F 2a FOR INDUCTION OF THERAPEUTIC ABORTION E . Granström, R . Gréen, M . Bygdeman, M . Toppozada .and N . Wiqvist (Received in final form 30 December 1972) Department of Chemistry, Rarolinska Institutet, S-104 01 Stockholm 60, Sweden and the Department of Obstetrics and Gynaecology, Rarolinska Sjukhuset, S-104 01 Stockholm 60, Sweden . SUMMARY Prostaglandin F2 , unlabeled or tritium-labeled, was administered intraamnioticaii~y to eight women in the second trimester of pregnancy . The absorption of PGF2 from amniotic fluid and the metabolism of the compound were sttiffied in samples of blood, amniotic fluid, and urine . The radioactivity in amniotic fluid was found to disappear very slowly, and 5 to 10 hrs after the administration several other radioactive compounds appeared in the amniotic fluid . The main product was identified as 9a,lla-dihydroxy-15-ketoprost-5-enoic acid by gas chromatography-mass spectrometry of two derivatives . The metabolites excreted into urine were found to be identical with those obtained after intravenous injection of PGF . Low concentrations of PGF , around 0 .2-0 .3 nanog/ml, were fouRa in plasma following intraaAgiotic administration . The metabolite was found in much higher concentrations ; the maximum levels ranged between 1 .5 and 14 .8 nanog/ml . In recent years prostaglandins have been successfully employed for termination of pregnancy . In the earlier investigations, the prostaglandins, generally PGF2a or PGE2 1) , were administered by intravenous infusion, but later the vaginal, extraamniotic or intraamniotic routes of administration have been used (1-6) . 1) The following trivial names and abbreviations are used : PGF (prostaglandin F ), 9a,lla,15-trihydroxyprosta-5,13-dienoic aRd ; PGE (prostaglan&Tn E), lla,15-dihydroxy-9-ketoprosta-5,13-dienoic acid ; 15-keto-dihydro-PGF2a , 9a,lla-dihydroxy-15-ketoprost-5-enoic acid . 219
280
Metabolism of PGF2a
Vol . 12, No. 5
When given intravenously PGF 2a is rapidly converted mainly into 9a,lla-dihydroxy-15-ketoprost-5-enoic acid and 9a,lla,15-trihydroxyprost-5-enoic acid (7) . Methods for quantitative determina tions of picomole amounts of PGF2a and these two metabolites have recently been developed in our laboratory (8,9) . Utilizing these methods the blood levels of the three compounds during and after intravenous administration of PGF 2a have been determined in a number of cases (10,11) . The present report deals with intraamniotic administration of PGF2a ; some kinetic and metabolic data as well as blood levels of the compound and one metabolite are reported .
EXPERIMENTAL PROCEDURE Five women of various gestational ages
(16-23 weeks, cases 1-
5) were given PGF2a intraamniotically . Each subject received 25 mg of
[9ß- 3H1-PGF 2a (32 pCi)
in a single injection . Samples of amnio-
tic fluid were collected at intervals, about 1 hr, 2, 5, 10 and (in one case) 20 hrs after the administration . The first portion of the sample withdrawn through the catheter was discarded . Urine was collected for 24 hrs after the injection in 6 hr portions . The radioactivity of aliquots of the amniotic fluid samples was measured in a Packard Tri-Carb Liquid Scintillation Spectrometer, model 3375 . The scintillation fluid consisted of 100 ml of Bio-Sole and 42 ml of Liquifluor in 1000 ml of toluene . Certain samples were acidified to pH 3 with 2 N HC1 and extracted either with ether or using Amberlite XAD-2 (12) . The extracts were subjected to reversed phase partition chromatography, first in solvent system C-50 (7) and subsequently after esterification of material in radioactive peaks with diazomethane, in solvent system F-55
(7) . Products
isolated using this procedure were converted into trimethylsilyl
Vol. 12, No . 5
Metabolism of PGF2a
221
ether derivatives or acetates and analyzed by gas chromatography (Barber--Colman gas chromatograph, series 5000) and mass spectrometry in combination with gas chromatography (LRB 9000 instrument) . The stationary phase was 18 OV-1 on Chromosorb W . Urinary samples were assayed for radioactivity as described above . The samples were combined, acidified to pH 3 and extracted using Amberlite XAD-2 . The extract was purified using silicic acid chromatography and reversed phase partition chromatography as described in detail elsewhere (12) . Peripheral venous blood (20 ml) was collected in heparinized tubes before and every sixth hour after intraamniotic administration of 25 mg of PGF2a to five midpregnant women (cases 4-8) . Two patients also received C9ß-3HJ-PGF2a (32 pCi, case 4 and 5, cf . above) . Plasma was immediately isolated and kept frozen until assayed . About 5 Ug of of
3,3,4,4-D4
1-
-17,18-3 H2,-PGF2a and about 1 ug 4J J-9a,lla-dihydroxy -15 - ketoprost-5 -enoic
[3,3,4,4-D
C17,18- 3H2
acid were added to each plasma sample and the amounts of corresponding non-deuterated compounds were determined with the gas chromatographic-mass spectrometric method described elsewhere (8) . This technique and the amounts of carriers used allow for determination of a total amount in the plasma samples of 10 nanog of PGF 2a and 1 nanog of 9a,lla-dihydroxy-15-ketoprost-5-enoic acid with accuracies of +7 .48 (SD) and 178 (SD), respectively . RESULTS Some data from the five experiments with 19a- HJ-PGF 2a are 3 presented in Table i . Analysis of the amniotic fluid samples revealed a slow disappearance of radioactivity from the amniotic sac, roughly inversely proportional to the amniotic fluid volume (Fig .l) .
222
Metabolism of PGF2 ac
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Vol . 12, No. 5
Metabolism of PGF2 at
223
FIG . 1 Disappearance of radioactivity from the amniotic fluid of five subjects . Some data from the five experiments are given in Table I .
In two cases the initial values were considerably lower than the subsequent ones (cases 3 and 5) which may be due to incomplete
mixing of the compound with the amniotic fluid . The amniotic fluid volumes were calculated from the disappearance curves by extrapolation to zero time and knowledge of the given dose .
In four cases (2-5) the samples obtained 6-10 hrs after the administration were analyzed for contents of metabolites . Different degrees of metabolism were found in the four cases . Fig . 2 shows the reversed phase partition chromatogram of an ether extract of
the 10 hr sample from case 2 . Essentially all of the radioactivity had been recovered in the ether extraction, and about 758 of this radioactivity were found in peak I (PGF 2., see below), whereas the remaining 258 were found in peaks appearing later in the chromatogram . In the three other cases (3-5) the administered PGF2. had been metabolized to 158, 128 and 28, respectively .
224
Vol. 12, No . 5
Metabolism of PGF2 0C
FIG . 2 Reversed phase partition chromatogram of an ether extract of amniotic fluid . The sample (10 ml) was obtained from patient G-B J (case 2) 10 hrs after the administration of 25 mg of PGF (32 yCi) . Solvent system : C-50 ; column : 4 .5 g of hydro-phobiii"Hyflo Super-Cel ; fraction volume : 5 ml ; 0 .2 ml were assayed for radioactivity .
10
20 30 Fraction number
i0
The identity of the material in peak I with PGF2a was established by gas chromatography-mass spectrometry of the methyl ester and trimethylsilyl ether . The material in peak II (Fig . 2) was esterified with diazomethane . On reversed phase partition chromatography in solvent system F-55 the material was found to be heterogenous and appeared as several peaks of radioactivity . Due to the small amount of material these compounds were not studied further . Peak III had an elution volume similar to that of 9a,lla-dihydroxy-15-ketoprost-5-enoic acid (120-150 ml on a 4 .5 g column (7)) . After esterification with diazomethane the compound was eluted as one single peak with an elution- volume of 80-110 ml (4 .5 g column, solvent F-55) . The material in this peak was converted into the trimethylsilyl ether or acetate (12) and was analyzed by gas chromatography-mass spectrometry . The data obtained on this analysis proved the identity of the compound with 9a,lla-dihydroxy-15-ketoprost-5-enoic acid (13) .
Vol. 12, No . 5
Metabolism of PGF2 ac
225
The excretion of radioactive products into urine in 24 hrs ranged between 8 and 248 of the given dose
(Table I) . Case 4, which
showed a more rapid disappearance of radioactivity from the amnio tic sac also excreted the metabolites more rapidly into urine (438 in 18 hrs) . The urinary portions were pooled for each patient and processed as described in detail elsewhere (12) . Analysis of the urinary metabolites showed exactly the same pattern of products as that obtained after intravenous administration (12,14), the main metabolite being 5a,7a-dihydroxy-11-ketotetranorprosta-1,16-dioic acid . The plasma levels of PGF2a and 9a,lla-dihydroxy-15-ketoprost-5-enoic acid are shown in Table II together with some clinical data . The concentration of PGF2a after administration were low in all cases, usually around 0 .2-0 .3 nanog/ml . The maximum levels of the metabolite ranged from 1 .48 nanog/ml to 14 .8 nanog/ml plasma . In cases number 5 and 6 a continuous rise of the concentration was seen during the time of study whereas in the other cases a maximum value was reached followed by a decline .
DISCUSSION This study demonstrates that prostaglandin F2c disappears slowly from the amniotic fluid following intraamniotic administration . With time a number of PGF 2a metabolites appear in the amnio tic fluid . The main metabolite was identified as 9a,lla-dihydroxy-15-ketoprost-5-enoic acid . Similar findings were recently reported by Pace-Asciak et al . (15) . The origin of the metabolite is not clear . The enzymes responsible for the transformations leading to this product,*i .e . 15hydroxyprostanoate dehydrogenase and prostaglandin-A 13 -reductase have been shown to be widely distributed in various tissues (16) .
226
Vol . 12, No. 5
Metabolism of PGF2 as
TABLE II Peripheral plasma levels of prostaglandin F and 9a,lla-dihydroxy-15-ketoprost-5-enoic acid . All times are rhative to the intraamniotic administration of 25 mg of prostaglandin F2a' Patient
Time
PGF ng/4Y
15-keto-dihydro-PGF 2a ng/ml
LA (case 4) (16 weeks)
0 6 12 18
0 .1 0 .3 0 .1
0 .60 14 .80 0 .0
ER (case 5) (17 weeks)
0 1 2 5 9
0 .1 0 .3 0 .4 0 .3 <0 .4
2 .15 3 .68 4 .98 5 .51
0 6 12 18
0 .0 0 .4 0 .3 0 .4
0 .04 2 .90 2 .09 3 .46
RMR (case 0 7) (20 6 weeks) 12 18
0 .0 0 .3 0 .3 0 .1
0 .04 1 .48 0 .80 0 .16
0 .3 0 .2 0 .2 0 .2
3 .00 3 .52 3 .75 3 .35
OF (case 6) (20 weeks)
MH (case 8) (17 weeks)
0 6 12 18 24
Inductionabortion interval hrs
Time of vomiting (no diarrhoea occurred) 6,7,9
15
115 10 330 301 275
21 10,10
42
0 603 (392 time 10) 0 51 165
25
20 .5
Mean uterine activity (M .U .)
0 244 (242 time 10) 0 301 270 208 (206 time 21)
Recently, it was demonstrated that the human placenta at term contains large amounts of 15-hydroxyprostanoate dehydrogenase (17) . Thus, it is likely that 9a,lla-dihydroxy-15-ketoprost-5-enoic acid is formed from the administered PGF 2a by enzymes in placental or fetal tissues . The plasma levels of prostaglandin F 2a found after intraamniotic instillation (0 .1-0 .4 ng/ml) indicate that minor amounts of this compound can pass from the amniotic sac into the maternal cir-
Vol. 12, No. 5
227
Metabolism of PGF 20C
culation . The values are considerably lower than those found earlier during intravenous administration of prostaglandin F 2a
(75
jig/min for 10 hrs) using the same methods for quantitation (1 .35 .7 ng/ml)
(11) . In the present study the plasma concentration of
the metabolite, 9a,lla-dihydroxy-15-ketoprost-5-enoic acid, reached much lower values than those obtained .during intravenous infusion of prostaglandin F 2a at a constant rate of 75 jig/min (10,11) . In those studies the ratio between the level of 9a,lla-dihydroxy-15-ketoprost-5-enoic acid and that of prostaglandin F2a was found to range from 9 .7 to 73 .5 . Kinetic studies have demonstrated that prostaglandin F2a in the systemic circulation is rapidly transformed into this metabolite (7) . The level of 9a,lla-dihydroxy-15-ketoprost-5-enoic acid can therefore be used as a sensitive indicator for release of prostaglandin F2a into the blood stream . However, the plasma levels of the metabolite found in the present study might not only reflect formation from prostaglandin F2a appearing in the blood but also a direct release of the metabolite from the amniotic fluid to the systemic circulation . The very low levels of PGF2a in peripheral plasma and the low incidence of side effects in relation to the induced uterine activity indicate that after intraamniotic administration, prostaglandin F2a exerts a local effect on the myometrium . It is reasonable to assume that the gastrointestinal side effects seen in case 4, 7 and 8 reflect some leakage of the injected dose from the amniotic fluid to the systemic circulation . However, such an explanation was not confirmed by the analyses of blood plasma, since the PGF 2a concentrations (0 .1-0 .4 nanog/ml) were far below the levels believed to be necessary to induce those side effects (>1 nanog/ml)
(10,
11) . It is also of interest to note that no significant amounts of
228
Vol. 12, No. 5
Metabolism of PGF2 of
9a,lla-dihydroxy-15-ketoprost-5-enoic acid could be detected in peripheral plasma 3 hrs after abortion had occurred (case 4) . ACKNOWLEDGEMENTS This project was supported by the Swedish Medical Research Council (proj .no . 13X-217 and 17X-2019), Stiftelsén Riksbankens Jubileumsfond and the World Health Organization . REFERENCES 1.
BYGDEMAN, M. and WIQVIST, N ., Ann . N .Y . Acad . Sci . l$2, 473 (1971) .
2.
KARIM, S .M .M ., Ann . N .Y . Acad . Sci . J$Q, 483 (1971) .
3.
EMBREY, M ., Ann . N .Y . Acad . Sci .
4.
WIQVIST, N ., BYGDEMAN, M . and TOPPOZADA, M ., Acta Obstet . Gynec . Scand . 22, 381 (1971) .
5.
TOPPOZADA, M ., BYGDEMAN, M . and WIQVIST, N ., Contraception 4, 293 (1971) .
6.
KARIM, S .M .M . and SHARMA, S .D ., Lancet 2, 47
7.
GRANSTROM, E ., Eur . J . Biochem . 22, 462 (1972) .
8.
GREEN, K ., GRANSTROM, E ., SAMUELSSON, B . and AXEN, U ., To be published . (1973) .
9.
GREEN, K ., GRANSTROM, E . and SAMUELSSON, B ., Third Conference on Prosta landins in Fertility Control, WHO Research and Training Centre on Human Reproduction, Karolinska Institutet, Stockholm (1972), p . 92 .
10 .
GREEN, K ., BEGUIN, F ., BYGDEMAN, M ., TOPPOZADA, M . and WIQVIST, N ., Ibid . p . 189 .
11 .
BEGUIN, F ., BYGDEMAN, M ., GREEN, K ., TOPPOZADA, M . and WIQVIST, N ., Acta Physiol . Scand . In press (1972) .
12 .
GRANSTROM, E . and SAMUELSSON, B ., J . Biol . Chem . _.16-, 7470 (1971) .
13 .
GRANSTR6M, E ., Eur . J . Biochem . 2Q, 451 (1972) .
14 .
GRANSTROM, E ., Eur . J . Biochem . 2R, 581 (1972) .
15 .
PACE-ASCIAK, C ., WOLFE, L .S ., GILLETT, P .G . and KINCH, R .A ., Prostaglandins 1, 469 (1972) .
518 (1971) .
(1971) .
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229
Metabolism of PGF 20C
16 .
KNGGARD, E ., LARSSON, C . and SAMUELSSON, B ., Acta Physiol . Scand . 21, 396 (1971) .
17 .
JARABAR, J ., Proc . Nat . Acad . Sci . USA
§2,
533 (1972) .
iâareV a tBruinart I'
PP . 219-229, 1973
Pergamon Press
METABOLIC AND QUANTITATIVE STUDIES IN CONNECTION WITH INTRAAMNIOTIC ADMINISTRATION OF PROSTAGLANDIN F 2a FOR INDUCTION OF THERAPEUTIC ABORTION E . Granström, R . Gréen, M . Bygdeman, M . Toppozada .and N . Wiqvist (Received in final form 30 December 1972) Department of Chemistry, Rarolinska Institutet, S-104 01 Stockholm 60, Sweden and the Department of Obstetrics and Gynaecology, Rarolinska Sjukhuset, S-104 01 Stockholm 60, Sweden . SUMMARY Prostaglandin F2 , unlabeled or tritium-labeled, was administered intraamnioticaii~y to eight women in the second trimester of pregnancy . The absorption of PGF2 from amniotic fluid and the metabolism of the compound were sttiffied in samples of blood, amniotic fluid, and urine . The radioactivity in amniotic fluid was found to disappear very slowly, and 5 to 10 hrs after the administration several other radioactive compounds appeared in the amniotic fluid . The main product was identified as 9a,lla-dihydroxy-15-ketoprost-5-enoic acid by gas chromatography-mass spectrometry of two derivatives . The metabolites excreted into urine were found to be identical with those obtained after intravenous injection of PGF . Low concentrations of PGF , around 0 .2-0 .3 nanog/ml, were fouRa in plasma following intraaAgiotic administration . The metabolite was found in much higher concentrations ; the maximum levels ranged between 1 .5 and 14 .8 nanog/ml . In recent years prostaglandins have been successfully employed for termination of pregnancy . In the earlier investigations, the prostaglandins, generally PGF2a or PGE2 1) , were administered by intravenous infusion, but later the vaginal, extraamniotic or intraamniotic routes of administration have been used (1-6) . 1) The following trivial names and abbreviations are used : PGF (prostaglandin F ), 9a,lla,15-trihydroxyprosta-5,13-dienoic aRd ; PGE (prostaglan&Tn E), lla,15-dihydroxy-9-ketoprosta-5,13-dienoic acid ; 15-keto-dihydro-PGF2a , 9a,lla-dihydroxy-15-ketoprost-5-enoic acid . 219
280
Metabolism of PGF2a
Vol . 12, No. 5
When given intravenously PGF 2a is rapidly converted mainly into 9a,lla-dihydroxy-15-ketoprost-5-enoic acid and 9a,lla,15-trihydroxyprost-5-enoic acid (7) . Methods for quantitative determina tions of picomole amounts of PGF2a and these two metabolites have recently been developed in our laboratory (8,9) . Utilizing these methods the blood levels of the three compounds during and after intravenous administration of PGF 2a have been determined in a number of cases (10,11) . The present report deals with intraamniotic administration of PGF2a ; some kinetic and metabolic data as well as blood levels of the compound and one metabolite are reported .
EXPERIMENTAL PROCEDURE Five women of various gestational ages
(16-23 weeks, cases 1-
5) were given PGF2a intraamniotically . Each subject received 25 mg of
[9ß- 3H1-PGF 2a (32 pCi)
in a single injection . Samples of amnio-
tic fluid were collected at intervals, about 1 hr, 2, 5, 10 and (in one case) 20 hrs after the administration . The first portion of the sample withdrawn through the catheter was discarded . Urine was collected for 24 hrs after the injection in 6 hr portions . The radioactivity of aliquots of the amniotic fluid samples was measured in a Packard Tri-Carb Liquid Scintillation Spectrometer, model 3375 . The scintillation fluid consisted of 100 ml of Bio-Sole and 42 ml of Liquifluor in 1000 ml of toluene . Certain samples were acidified to pH 3 with 2 N HC1 and extracted either with ether or using Amberlite XAD-2 (12) . The extracts were subjected to reversed phase partition chromatography, first in solvent system C-50 (7) and subsequently after esterification of material in radioactive peaks with diazomethane, in solvent system F-55
(7) . Products
isolated using this procedure were converted into trimethylsilyl
Vol. 12, No . 5
Metabolism of PGF2a
221
ether derivatives or acetates and analyzed by gas chromatography (Barber--Colman gas chromatograph, series 5000) and mass spectrometry in combination with gas chromatography (LRB 9000 instrument) . The stationary phase was 18 OV-1 on Chromosorb W . Urinary samples were assayed for radioactivity as described above . The samples were combined, acidified to pH 3 and extracted using Amberlite XAD-2 . The extract was purified using silicic acid chromatography and reversed phase partition chromatography as described in detail elsewhere (12) . Peripheral venous blood (20 ml) was collected in heparinized tubes before and every sixth hour after intraamniotic administration of 25 mg of PGF2a to five midpregnant women (cases 4-8) . Two patients also received C9ß-3HJ-PGF2a (32 pCi, case 4 and 5, cf . above) . Plasma was immediately isolated and kept frozen until assayed . About 5 Ug of of
3,3,4,4-D4
1-
-17,18-3 H2,-PGF2a and about 1 ug 4J J-9a,lla-dihydroxy -15 - ketoprost-5 -enoic
[3,3,4,4-D
C17,18- 3H2
acid were added to each plasma sample and the amounts of corresponding non-deuterated compounds were determined with the gas chromatographic-mass spectrometric method described elsewhere (8) . This technique and the amounts of carriers used allow for determination of a total amount in the plasma samples of 10 nanog of PGF 2a and 1 nanog of 9a,lla-dihydroxy-15-ketoprost-5-enoic acid with accuracies of +7 .48 (SD) and 178 (SD), respectively . RESULTS Some data from the five experiments with 19a- HJ-PGF 2a are 3 presented in Table i . Analysis of the amniotic fluid samples revealed a slow disappearance of radioactivity from the amniotic sac, roughly inversely proportional to the amniotic fluid volume (Fig .l) .
222
Metabolism of PGF2 ac
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Vol . 12, No. 5
Metabolism of PGF2 at
223
FIG . 1 Disappearance of radioactivity from the amniotic fluid of five subjects . Some data from the five experiments are given in Table I .
In two cases the initial values were considerably lower than the subsequent ones (cases 3 and 5) which may be due to incomplete
mixing of the compound with the amniotic fluid . The amniotic fluid volumes were calculated from the disappearance curves by extrapolation to zero time and knowledge of the given dose .
In four cases (2-5) the samples obtained 6-10 hrs after the administration were analyzed for contents of metabolites . Different degrees of metabolism were found in the four cases . Fig . 2 shows the reversed phase partition chromatogram of an ether extract of
the 10 hr sample from case 2 . Essentially all of the radioactivity had been recovered in the ether extraction, and about 758 of this radioactivity were found in peak I (PGF 2., see below), whereas the remaining 258 were found in peaks appearing later in the chromatogram . In the three other cases (3-5) the administered PGF2. had been metabolized to 158, 128 and 28, respectively .
224
Vol. 12, No . 5
Metabolism of PGF2 0C
FIG . 2 Reversed phase partition chromatogram of an ether extract of amniotic fluid . The sample (10 ml) was obtained from patient G-B J (case 2) 10 hrs after the administration of 25 mg of PGF (32 yCi) . Solvent system : C-50 ; column : 4 .5 g of hydro-phobiii"Hyflo Super-Cel ; fraction volume : 5 ml ; 0 .2 ml were assayed for radioactivity .
10
20 30 Fraction number
i0
The identity of the material in peak I with PGF2a was established by gas chromatography-mass spectrometry of the methyl ester and trimethylsilyl ether . The material in peak II (Fig . 2) was esterified with diazomethane . On reversed phase partition chromatography in solvent system F-55 the material was found to be heterogenous and appeared as several peaks of radioactivity . Due to the small amount of material these compounds were not studied further . Peak III had an elution volume similar to that of 9a,lla-dihydroxy-15-ketoprost-5-enoic acid (120-150 ml on a 4 .5 g column (7)) . After esterification with diazomethane the compound was eluted as one single peak with an elution- volume of 80-110 ml (4 .5 g column, solvent F-55) . The material in this peak was converted into the trimethylsilyl ether or acetate (12) and was analyzed by gas chromatography-mass spectrometry . The data obtained on this analysis proved the identity of the compound with 9a,lla-dihydroxy-15-ketoprost-5-enoic acid (13) .
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225
The excretion of radioactive products into urine in 24 hrs ranged between 8 and 248 of the given dose
(Table I) . Case 4, which
showed a more rapid disappearance of radioactivity from the amnio tic sac also excreted the metabolites more rapidly into urine (438 in 18 hrs) . The urinary portions were pooled for each patient and processed as described in detail elsewhere (12) . Analysis of the urinary metabolites showed exactly the same pattern of products as that obtained after intravenous administration (12,14), the main metabolite being 5a,7a-dihydroxy-11-ketotetranorprosta-1,16-dioic acid . The plasma levels of PGF2a and 9a,lla-dihydroxy-15-ketoprost-5-enoic acid are shown in Table II together with some clinical data . The concentration of PGF2a after administration were low in all cases, usually around 0 .2-0 .3 nanog/ml . The maximum levels of the metabolite ranged from 1 .48 nanog/ml to 14 .8 nanog/ml plasma . In cases number 5 and 6 a continuous rise of the concentration was seen during the time of study whereas in the other cases a maximum value was reached followed by a decline .
DISCUSSION This study demonstrates that prostaglandin F2c disappears slowly from the amniotic fluid following intraamniotic administration . With time a number of PGF 2a metabolites appear in the amnio tic fluid . The main metabolite was identified as 9a,lla-dihydroxy-15-ketoprost-5-enoic acid . Similar findings were recently reported by Pace-Asciak et al . (15) . The origin of the metabolite is not clear . The enzymes responsible for the transformations leading to this product,*i .e . 15hydroxyprostanoate dehydrogenase and prostaglandin-A 13 -reductase have been shown to be widely distributed in various tissues (16) .
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TABLE II Peripheral plasma levels of prostaglandin F and 9a,lla-dihydroxy-15-ketoprost-5-enoic acid . All times are rhative to the intraamniotic administration of 25 mg of prostaglandin F2a' Patient
Time
PGF ng/4Y
15-keto-dihydro-PGF 2a ng/ml
LA (case 4) (16 weeks)
0 6 12 18
0 .1 0 .3 0 .1
0 .60 14 .80 0 .0
ER (case 5) (17 weeks)
0 1 2 5 9
0 .1 0 .3 0 .4 0 .3 <0 .4
2 .15 3 .68 4 .98 5 .51
0 6 12 18
0 .0 0 .4 0 .3 0 .4
0 .04 2 .90 2 .09 3 .46
RMR (case 0 7) (20 6 weeks) 12 18
0 .0 0 .3 0 .3 0 .1
0 .04 1 .48 0 .80 0 .16
0 .3 0 .2 0 .2 0 .2
3 .00 3 .52 3 .75 3 .35
OF (case 6) (20 weeks)
MH (case 8) (17 weeks)
0 6 12 18 24
Inductionabortion interval hrs
Time of vomiting (no diarrhoea occurred) 6,7,9
15
115 10 330 301 275
21 10,10
42
0 603 (392 time 10) 0 51 165
25
20 .5
Mean uterine activity (M .U .)
0 244 (242 time 10) 0 301 270 208 (206 time 21)
Recently, it was demonstrated that the human placenta at term contains large amounts of 15-hydroxyprostanoate dehydrogenase (17) . Thus, it is likely that 9a,lla-dihydroxy-15-ketoprost-5-enoic acid is formed from the administered PGF 2a by enzymes in placental or fetal tissues . The plasma levels of prostaglandin F 2a found after intraamniotic instillation (0 .1-0 .4 ng/ml) indicate that minor amounts of this compound can pass from the amniotic sac into the maternal cir-
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227
Metabolism of PGF 20C
culation . The values are considerably lower than those found earlier during intravenous administration of prostaglandin F 2a
(75
jig/min for 10 hrs) using the same methods for quantitation (1 .35 .7 ng/ml)
(11) . In the present study the plasma concentration of
the metabolite, 9a,lla-dihydroxy-15-ketoprost-5-enoic acid, reached much lower values than those obtained .during intravenous infusion of prostaglandin F 2a at a constant rate of 75 jig/min (10,11) . In those studies the ratio between the level of 9a,lla-dihydroxy-15-ketoprost-5-enoic acid and that of prostaglandin F2a was found to range from 9 .7 to 73 .5 . Kinetic studies have demonstrated that prostaglandin F2a in the systemic circulation is rapidly transformed into this metabolite (7) . The level of 9a,lla-dihydroxy-15-ketoprost-5-enoic acid can therefore be used as a sensitive indicator for release of prostaglandin F2a into the blood stream . However, the plasma levels of the metabolite found in the present study might not only reflect formation from prostaglandin F2a appearing in the blood but also a direct release of the metabolite from the amniotic fluid to the systemic circulation . The very low levels of PGF2a in peripheral plasma and the low incidence of side effects in relation to the induced uterine activity indicate that after intraamniotic administration, prostaglandin F2a exerts a local effect on the myometrium . It is reasonable to assume that the gastrointestinal side effects seen in case 4, 7 and 8 reflect some leakage of the injected dose from the amniotic fluid to the systemic circulation . However, such an explanation was not confirmed by the analyses of blood plasma, since the PGF 2a concentrations (0 .1-0 .4 nanog/ml) were far below the levels believed to be necessary to induce those side effects (>1 nanog/ml)
(10,
11) . It is also of interest to note that no significant amounts of
228
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Metabolism of PGF2 of
9a,lla-dihydroxy-15-ketoprost-5-enoic acid could be detected in peripheral plasma 3 hrs after abortion had occurred (case 4) . ACKNOWLEDGEMENTS This project was supported by the Swedish Medical Research Council (proj .no . 13X-217 and 17X-2019), Stiftelsén Riksbankens Jubileumsfond and the World Health Organization . REFERENCES 1.
BYGDEMAN, M. and WIQVIST, N ., Ann . N .Y . Acad . Sci . l$2, 473 (1971) .
2.
KARIM, S .M .M ., Ann . N .Y . Acad . Sci . J$Q, 483 (1971) .
3.
EMBREY, M ., Ann . N .Y . Acad . Sci .
4.
WIQVIST, N ., BYGDEMAN, M . and TOPPOZADA, M ., Acta Obstet . Gynec . Scand . 22, 381 (1971) .
5.
TOPPOZADA, M ., BYGDEMAN, M . and WIQVIST, N ., Contraception 4, 293 (1971) .
6.
KARIM, S .M .M . and SHARMA, S .D ., Lancet 2, 47
7.
GRANSTROM, E ., Eur . J . Biochem . 22, 462 (1972) .
8.
GREEN, K ., GRANSTROM, E ., SAMUELSSON, B . and AXEN, U ., To be published . (1973) .
9.
GREEN, K ., GRANSTROM, E . and SAMUELSSON, B ., Third Conference on Prosta landins in Fertility Control, WHO Research and Training Centre on Human Reproduction, Karolinska Institutet, Stockholm (1972), p . 92 .
10 .
GREEN, K ., BEGUIN, F ., BYGDEMAN, M ., TOPPOZADA, M . and WIQVIST, N ., Ibid . p . 189 .
11 .
BEGUIN, F ., BYGDEMAN, M ., GREEN, K ., TOPPOZADA, M . and WIQVIST, N ., Acta Physiol . Scand . In press (1972) .
12 .
GRANSTROM, E . and SAMUELSSON, B ., J . Biol . Chem . _.16-, 7470 (1971) .
13 .
GRANSTR6M, E ., Eur . J . Biochem . 2Q, 451 (1972) .
14 .
GRANSTROM, E ., Eur . J . Biochem . 2R, 581 (1972) .
15 .
PACE-ASCIAK, C ., WOLFE, L .S ., GILLETT, P .G . and KINCH, R .A ., Prostaglandins 1, 469 (1972) .
518 (1971) .
(1971) .
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Metabolism of PGF 20C
16 .
KNGGARD, E ., LARSSON, C . and SAMUELSSON, B ., Acta Physiol . Scand . 21, 396 (1971) .
17 .
JARABAR, J ., Proc . Nat . Acad . Sci . USA
§2,
533 (1972) .