ACTA ENDOCRINOLOGICA
83
University
(1976)
410-419
Department of Clinical Chemistry, of Helsinki, Meilahti Hospital, SF-00290 Helsinki 29
OESTROGEN IN HUMAN PREGNANCY FAECES
By Herman Adlercreutz and Finian Martin1)
ABSTRACT The oestrogen content of two 24 h pools of pregnancy faeces, obtained from 2 normal women in the 33rd\p=n-\37th week og gestation, was studied. The qualitative analyses were made by gas chromatography mass spectrometry and the quantitative analyses by mass fragmentography. The presence of the following oestrogens in pregnancy faeces was established: Oestriol, oestrone, oestradiol-17\g=b\, 16-epioestriol, 17-epioestriol, 16\g=a\-hydroxyoestrone, 16-oxo-oestradiol-17\g=b\, 15\g=a\-hydroxyoestrone and 15\g=a\-hydroxyoestradiol-17\g=b\. In addition, mass fragmentographic evidence was obtained for the presence of 16\g=b\-hydroxyoestrone, 2-methoxyoestrone and oestradiol-17\g=a\. The total oestrogen excretion determined in the two pools was 786 and 1300 \g=m\g per 24 h. Unconjugated oestrogens accounted for 97.8 and 98.6% of these amounts, respectively. Oestriol, oestradiol-17\g=b\, 15\g=a\-hydroxyoestradiol-17\g=b\,16-epioestriol and oestrone, in that order, were quantitatively the most significant of the oestrogens determined. The remarkably high levels of oestradiol-17\g=b\ found in faeces show, that in pregnancy, this mode of excretion is as important as urine for the elimination of this biologically active steroid. It is suggested that some of the oestradiol may have been formed through bacterial enzyme action from other oestrogens or neutral steroids. Only trace amounts of ring D \g=a\-ketolic oestrogens were found in faeces. This is in marked contrast to the considerable amounts of these steroids found in pregnancy bile and urine. -
0 Present address: of Ireland.
Department
of
Biochemistry, Trinity College,
Dublin 2.
Republic
The presence of oestrogen activity in human faeces was observed as early as 1928-1930 (Dohrn & Faure 1928; Luchsinger & Voss 1929; Siebke & Schuschania 1930). Much later, Sandberg & Slaunwhite (1957, 1965) demonstrated that the intravenous administration of [14C] oestrone, [14C] oestradiol and [14C]\x=req-\ oestriol to normal female subjects resulted in a maximal excretion of 4.4\m=-\ 18.0% of the administered radioactivity in the faeces. Considerable amounts of oestrogens are secreted in human bile during pregnancy, all of which are present in conjugated form (Adlercreutz 1962; Adlercreutz et al. 1973). The fate of these steroids in the intestinal tract and the nature and extent of their eventual faecal excretion has not been documented. In this report the identification and quantitation of 12 endogenous oestrogens in late pregnancy faeces is described. The pattern obtained is compared to that found in plasma, bile and urine and the role of the intestinal tract in oestrogen metabolism in pregnancy is discussed.
MATERIAL AND METHODS
Faeces was collected from 2 normal women, in the 33rd-37th week of gestation, for two consecutive 24 h periods (day 1 and 2). The faeces was frozen and stored at -18°C until analyzed.
Steroids
OEj (oestrone): 3-hydroxy-l,3,5(10)-oestratrien-17-one; OE2 (oestradiol): l,3,5(10)-oestratriene-3,17/?-diol; 0E3 (oestriol): l,3,5(10)-oestratriene-3,16a,17/?-triol; OE,-17a (oestradiol-17a): l,3,5(10)-oestratriene-3,17a-diol; 2-MeOEj (2-methoxyoestrone): 3-hydroxy-l,3,5(10)-oestratrien-17-one-2-methyl ether; 16-cpiOE3 (16-epioestriol): 1,3,5(10)-oestratriene-3,16/5,17/S-triol; 17-epiOE:i (17-epioestriol): l,3,5(10)-oestratriene-3,16a,17a-triol; 16,17-epiOE3 (16,17-epioestriol): l,3,5(10)-oestratriene-3,16/3,17a-triol; 16a-OHOE, (16a-hydroxyoestrone): 3,16a-dihydroxy-l,3,5(10)-oestratrien-17-one; 16/J-OHOE, (16/?-hydroxyoestrone): 3,16/>-dihydroxy-l,3,5(10)-oestratrien-17-one; 16-oxoOE.> (16-oxo-oestradiol): 3,17/?-dihydroxy-l,3,5(10)-oestratrien-16-one; 15a-OHOEj (15a-hydroxyoestrone): 3,15a-dihydroxy-l,3,5(10)-oestratrien-17-one; 15a-OHOEo (I5a-hydroxyoestradiol): l,3,5(10)oestratriene-3,15a,17/?-triol. Extraction of faeces and
preparation of pools
Prior to extraction the frozen faeces samples were left at 4°C overnight to thaw. The samples were then weighed and homogenized in acetone-ethanol (1:1, v/v) 10 volumes/sample weight (e. g. sample weight 200 g, solvent volume 2 1) using a Sorvall Omnimix at full speed for 10 min. The homogenate was filtered under pressure and the residue washed with the same volume of acetone-ethanol (1:1) and filtered. The resulting residue was subjected to the same procedure a second time but this time
using chloroform-methanol (1:1, v/v) as solvent. All filtrates were then combined and evaporated to dryness. The extracts were stored in water-methanol (3:7, v/v) at -18°C. For analysis the day 1 extracts from the 2 patients were pooled, as were the day 2 extracts. Both pools were evaporated to dryness and then, dissolved in 100 ml of chloroform-methanol (1:1). For quantitative analysis 5 per cent of each pool was taken, dried, dissolved in a minimal volume of water-methanol (3:7, v/v) and left at -18°C for 48 h. The pools were then centrifuged at -20°C and the re¬ sulting supernatant taken for analysis. To obtain mass spectra larger amounts of starting material were used.
Preparation of faecal (GC-MS)
extracts
for
gas
chromatography
—
mass
spectrometry
This was done as described for bile by Adlercreutz Se Luukkainen (1967, 1968) with slight modifications (Adlercreutz 1975). Unconjugated oestrogens were extracted with ethyl ether from the faceal extract dissolved in urate buffer as described previously for plasma oestrogens (Adlercreutz 1975). For the first time 15a-OHOE2 was identified and quantitated in the OE3 fraction. Procedural losses for this compound were cor¬ rected for
on
the basis of
Identification by
gas
OE3
recovery.
chromatography
mass
spectrometry
LKB 9000 Gas Chromatograph mass spectrometer (LKB-Produkter AB, Bromma, Sweden) using the conditions described in Adlercreutz et al. (1973) except that 3% OV-210 columns were used instead of QF-1. The criteria used for establishing the identity of the unknown oestrogens with the reference standards and the sources of the reference mass spectra of all the oestrogens considered in this report are given in Adlercreutz et al. (1973). Some of the oestrogen fractions contained such small amounts of one or several oestrogens that mass spectra could not be obtained. In these cases Chromatographie and mass fragmentographic evidence for identity was obtained by monitoring the molecular ion and other prominent or characteristic ions during GC-MS. If their behaviour was identical with those of the reference standards the steroids were re¬ garded as identified with reservation. In this connection it must be emphasized that mass spectra were, in most cases, obtained from the unconjugated fractions. Therefore, if identical mass fragmentographic and Chromatographie behaviour as standards was obtained for the same compounds in the conjugated fraction, this was regarded as satisfactory evidence for their presence in this fraction, also.
Mass spectra
-
were
recorded
on an
-
Quantitation by
mass
fragmentography
Quantitative analysis of the oestrogens in the faeces samples was carried out by mass fragmentography. The conditions used were almost exactly as described by Adlercreutz et al. (19745) for plasma and biliary measurements. Some measurements were also made on a new LKB 2091 gas chromatography mass spectrometer in the LKB applications laboratory in Stockholm. In the LKB 9000 GC-MS instrument we used a 3 "la OV-210 column instead of the 1 % QF-1 column. Only peak heights were measured and not peak areas as in the previous report. Values were corrected for losses incurred during purification on the basis of simultaneous recovery experiments using bile samples, as described earlier (Adlercreutz et al. 19745). -
RESULTS were identified in the unconjugated fraction of preg¬ faeces the basis on of mass spectra identical with those of reference nancy standards: OE3, OE,, OE2, 16a-OHOE1; 16-oxoOE2, 15a-OHOE1; 15a-OHOE2, 16-epiOE3 and 17-epiOE3. 16/5-OHOEi was identified with reservation on the basis of a slightly contaminated mass spectrum and by mass fragmentography. In addition, 2-MeOEj and OE2-17a were detected in the unconjugated fraction by mass fragmentography, only. The mass spectra obtained for the derivative of OE2 is shown in Fig. 1, and in Fig. 2 the mass spectrum of the trimethylsilyl ether (TMS) derivative of 15a-OHOE2 is compared to that of the correspond¬ ing reference standard. 16,17-epiOE3-TMS (Fig. 3) has a retention time close to that of 15a-OHOE2-TMS on OV-210 and slight contamination due to this compound could be observed in the mass spectrum obtained for 15a-OHOE->TMS (Fig. 2). OE3, OEi and 15a-OHOE2 in the conjugated fraction were identified on the basis of mass spectra which were identical with those of the corresponding reference standards. In addition, the following oestrogens in the conjugated fraction were detected by mass fragmentography: 2-MeOEi, OE2-17/5, 16epiOE3, 16a-OHOE,, 16/Î-OHOE,, 16-oxoOE2 and 15a-OHOE!. In Table 1 the concentrations of the 12 oestrogens identified and quantitated in both the unconjugated and conjugated fractions of the two pools of preg¬ nancy faeces are given. In Table 2, the mean total daily faecal excretion of
The
following oestrogens
) 00
M
1227
-
1358
BO
60 10 i ri
20
160
I.
J
1S6
I| 240
211
Jk
•ft^i"., 100 M/E
ID-CODE
.
€1
051171
MOLECULE WEIGHT t 358. D 1. 0 7. PEAK THRESHOLD i ACC. VOLTAGE . 3.5 KV ELECTRON ENERGIES , 22. 5 EV (11) 2D. 0 EV (1) -
DEPT.
COLUMN
:
SE-30
TEMPERATURES
i
FLASH HEATER 230 C COLUMN 215 C SEPARATOR 250 C -
REL.
OF CLINICAL CHEMISTRY.
Fig.
RET.
-
TIME
:
.59
( CHOLESTANE -
UNIVERSITY OF HELSINKI
1.
Mass spectrum of the methylated and TMS derivative of oestradiol isolated from the unconjugated fraction of human pregnancy faeces.
100
M
;504 BO -
60 -
10
i
-
20 -
,
298 324
Jlr'.^rr-i't'ifAri.lfki M,',
II J4D 345_
270
i
(¡,4 I ,1. I\'>
r>r
^O" ,
X
j
>i
489
-r
r-r4-,
M/E
218 ID-CODE i 021071 MOLECULE WEIGHT 5 01. 0 PEAK THRESHOLD i 1.07. ACC. VOLTAGE : 3. 5 KV ELECTRON ENERGIES ( 20. 0 EV (I) 22. 5 EV (¡I)
COLUMN
DEPT.
i
OV-210
TEMPERATURES
-
FLASH HEATER 230 C COLUMN 220 C SEPARATOR 210 C 1.20 (CHOLESTANE TIME < .
-
-
PEL.
RET.
-
UNIVERSITY OF HELSINKI
OF CLINICAL CHEMISTRY.
100
M
504
80
.-rriU.iiy.,.,.L mwÍ^.U V
399
*
386, 414
lr\i i M/E
120 ID-CODE 010675 MOLECULE WEIGHT .501. 0 PEAK THRESHOLD i 1. 0 7.
COLUMN
ELECTRON ENERGIES = 20. 0 EV (I) 22. 5 EV
REL.
-
ACC.
VOLTAGE
DEPT.
3. 5 KV
:
OV-210
TEMPERATURES
FLASH HEATER 218 C COLUMN 208 C SEPARATOR 216 C TIME 1.23 ( CHOLESTANE .
-
-
til)
OF CLINICAL CHEMISTRY,
Pig.
RET.
-
UNIVERSITY OF HELSINKI
2.
Mass spectrum of the TMS derivative of 15a-hydroxyoestradiol fraction of human pregnancy faeces (top panel) and the corresponding reference standard (lower panel). The compound from faeces is slightly contaminated with 16,17-epioestriol, the mass spectrum of which is seen in Fig. 3.
11 of the
oestrogens is compared
to their
plasma, biliary and urinary concentrations in the last trimester of pregnancy (Adlercreutz et al. \974a,b). The total oestrogen excretion measured was 786 fig/24 h in the day 1 pool and 1300 /,g/24 h in the day 2 pool. Of this 97.8 and 98.6%, respectively, were present in the unconjugated fraction. The only oestrogen preferentially mean
M
100
504 BO 310
60
345
386
I
324
414
360
I.J...U II jJaMX M/E
ID-CODE . 010875 85 MOLECULE WEIGHT .501.0 PEAK THRESHOLD 2. 0 X ACC. VOLTAGE : 3. 5 KV ELECTRON ENERGIES . 20. 0 EV (I) 22. 5 EV (II)
COLUMN
DEPT.
;
OV-210
FLASH HEATER 21B C COLUMN 208 C SEPARATOR 218 C TIME i 1.31 (CHOLESTANE
TEMPERATURES
-
i
-
REL.
OF CLINICAL CHEMISTRY.
RET.
-
1
-
UNIVERSITY OF HELSINKI
Pig.
3.
Mass spectrum of the TMS derivative of
16,17-epioestriol reference
standard.
Table 1.
Oestrogen
concentrations in the
unconjugated
pools of human
and conjugated fractions of two 24-h pregnancy faeces.
Unconjugated Day
Conjugated
Day
1
Day
OE,
2-MeOE, OE2-17/? OE2-17a 16-epiOE3 17-epiOE3 16a-OHOE, 16^-OHOE,
160.5
244.5
0.4
3.6 90.3 22.9 2.8
6.0 194.0 53.6 3.0
83
University
(1976)
410-419
Department of Clinical Chemistry, of Helsinki, Meilahti Hospital, SF-00290 Helsinki 29
OESTROGEN IN HUMAN PREGNANCY FAECES
By Herman Adlercreutz and Finian Martin1)
ABSTRACT The oestrogen content of two 24 h pools of pregnancy faeces, obtained from 2 normal women in the 33rd\p=n-\37th week og gestation, was studied. The qualitative analyses were made by gas chromatography mass spectrometry and the quantitative analyses by mass fragmentography. The presence of the following oestrogens in pregnancy faeces was established: Oestriol, oestrone, oestradiol-17\g=b\, 16-epioestriol, 17-epioestriol, 16\g=a\-hydroxyoestrone, 16-oxo-oestradiol-17\g=b\, 15\g=a\-hydroxyoestrone and 15\g=a\-hydroxyoestradiol-17\g=b\. In addition, mass fragmentographic evidence was obtained for the presence of 16\g=b\-hydroxyoestrone, 2-methoxyoestrone and oestradiol-17\g=a\. The total oestrogen excretion determined in the two pools was 786 and 1300 \g=m\g per 24 h. Unconjugated oestrogens accounted for 97.8 and 98.6% of these amounts, respectively. Oestriol, oestradiol-17\g=b\, 15\g=a\-hydroxyoestradiol-17\g=b\,16-epioestriol and oestrone, in that order, were quantitatively the most significant of the oestrogens determined. The remarkably high levels of oestradiol-17\g=b\ found in faeces show, that in pregnancy, this mode of excretion is as important as urine for the elimination of this biologically active steroid. It is suggested that some of the oestradiol may have been formed through bacterial enzyme action from other oestrogens or neutral steroids. Only trace amounts of ring D \g=a\-ketolic oestrogens were found in faeces. This is in marked contrast to the considerable amounts of these steroids found in pregnancy bile and urine. -
0 Present address: of Ireland.
Department
of
Biochemistry, Trinity College,
Dublin 2.
Republic
The presence of oestrogen activity in human faeces was observed as early as 1928-1930 (Dohrn & Faure 1928; Luchsinger & Voss 1929; Siebke & Schuschania 1930). Much later, Sandberg & Slaunwhite (1957, 1965) demonstrated that the intravenous administration of [14C] oestrone, [14C] oestradiol and [14C]\x=req-\ oestriol to normal female subjects resulted in a maximal excretion of 4.4\m=-\ 18.0% of the administered radioactivity in the faeces. Considerable amounts of oestrogens are secreted in human bile during pregnancy, all of which are present in conjugated form (Adlercreutz 1962; Adlercreutz et al. 1973). The fate of these steroids in the intestinal tract and the nature and extent of their eventual faecal excretion has not been documented. In this report the identification and quantitation of 12 endogenous oestrogens in late pregnancy faeces is described. The pattern obtained is compared to that found in plasma, bile and urine and the role of the intestinal tract in oestrogen metabolism in pregnancy is discussed.
MATERIAL AND METHODS
Faeces was collected from 2 normal women, in the 33rd-37th week of gestation, for two consecutive 24 h periods (day 1 and 2). The faeces was frozen and stored at -18°C until analyzed.
Steroids
OEj (oestrone): 3-hydroxy-l,3,5(10)-oestratrien-17-one; OE2 (oestradiol): l,3,5(10)-oestratriene-3,17/?-diol; 0E3 (oestriol): l,3,5(10)-oestratriene-3,16a,17/?-triol; OE,-17a (oestradiol-17a): l,3,5(10)-oestratriene-3,17a-diol; 2-MeOEj (2-methoxyoestrone): 3-hydroxy-l,3,5(10)-oestratrien-17-one-2-methyl ether; 16-cpiOE3 (16-epioestriol): 1,3,5(10)-oestratriene-3,16/5,17/S-triol; 17-epiOE:i (17-epioestriol): l,3,5(10)-oestratriene-3,16a,17a-triol; 16,17-epiOE3 (16,17-epioestriol): l,3,5(10)-oestratriene-3,16/3,17a-triol; 16a-OHOE, (16a-hydroxyoestrone): 3,16a-dihydroxy-l,3,5(10)-oestratrien-17-one; 16/J-OHOE, (16/?-hydroxyoestrone): 3,16/>-dihydroxy-l,3,5(10)-oestratrien-17-one; 16-oxoOE.> (16-oxo-oestradiol): 3,17/?-dihydroxy-l,3,5(10)-oestratrien-16-one; 15a-OHOEj (15a-hydroxyoestrone): 3,15a-dihydroxy-l,3,5(10)-oestratrien-17-one; 15a-OHOEo (I5a-hydroxyoestradiol): l,3,5(10)oestratriene-3,15a,17/?-triol. Extraction of faeces and
preparation of pools
Prior to extraction the frozen faeces samples were left at 4°C overnight to thaw. The samples were then weighed and homogenized in acetone-ethanol (1:1, v/v) 10 volumes/sample weight (e. g. sample weight 200 g, solvent volume 2 1) using a Sorvall Omnimix at full speed for 10 min. The homogenate was filtered under pressure and the residue washed with the same volume of acetone-ethanol (1:1) and filtered. The resulting residue was subjected to the same procedure a second time but this time
using chloroform-methanol (1:1, v/v) as solvent. All filtrates were then combined and evaporated to dryness. The extracts were stored in water-methanol (3:7, v/v) at -18°C. For analysis the day 1 extracts from the 2 patients were pooled, as were the day 2 extracts. Both pools were evaporated to dryness and then, dissolved in 100 ml of chloroform-methanol (1:1). For quantitative analysis 5 per cent of each pool was taken, dried, dissolved in a minimal volume of water-methanol (3:7, v/v) and left at -18°C for 48 h. The pools were then centrifuged at -20°C and the re¬ sulting supernatant taken for analysis. To obtain mass spectra larger amounts of starting material were used.
Preparation of faecal (GC-MS)
extracts
for
gas
chromatography
—
mass
spectrometry
This was done as described for bile by Adlercreutz Se Luukkainen (1967, 1968) with slight modifications (Adlercreutz 1975). Unconjugated oestrogens were extracted with ethyl ether from the faceal extract dissolved in urate buffer as described previously for plasma oestrogens (Adlercreutz 1975). For the first time 15a-OHOE2 was identified and quantitated in the OE3 fraction. Procedural losses for this compound were cor¬ rected for
on
the basis of
Identification by
gas
OE3
recovery.
chromatography
mass
spectrometry
LKB 9000 Gas Chromatograph mass spectrometer (LKB-Produkter AB, Bromma, Sweden) using the conditions described in Adlercreutz et al. (1973) except that 3% OV-210 columns were used instead of QF-1. The criteria used for establishing the identity of the unknown oestrogens with the reference standards and the sources of the reference mass spectra of all the oestrogens considered in this report are given in Adlercreutz et al. (1973). Some of the oestrogen fractions contained such small amounts of one or several oestrogens that mass spectra could not be obtained. In these cases Chromatographie and mass fragmentographic evidence for identity was obtained by monitoring the molecular ion and other prominent or characteristic ions during GC-MS. If their behaviour was identical with those of the reference standards the steroids were re¬ garded as identified with reservation. In this connection it must be emphasized that mass spectra were, in most cases, obtained from the unconjugated fractions. Therefore, if identical mass fragmentographic and Chromatographie behaviour as standards was obtained for the same compounds in the conjugated fraction, this was regarded as satisfactory evidence for their presence in this fraction, also.
Mass spectra
-
were
recorded
on an
-
Quantitation by
mass
fragmentography
Quantitative analysis of the oestrogens in the faeces samples was carried out by mass fragmentography. The conditions used were almost exactly as described by Adlercreutz et al. (19745) for plasma and biliary measurements. Some measurements were also made on a new LKB 2091 gas chromatography mass spectrometer in the LKB applications laboratory in Stockholm. In the LKB 9000 GC-MS instrument we used a 3 "la OV-210 column instead of the 1 % QF-1 column. Only peak heights were measured and not peak areas as in the previous report. Values were corrected for losses incurred during purification on the basis of simultaneous recovery experiments using bile samples, as described earlier (Adlercreutz et al. 19745). -
RESULTS were identified in the unconjugated fraction of preg¬ faeces the basis on of mass spectra identical with those of reference nancy standards: OE3, OE,, OE2, 16a-OHOE1; 16-oxoOE2, 15a-OHOE1; 15a-OHOE2, 16-epiOE3 and 17-epiOE3. 16/5-OHOEi was identified with reservation on the basis of a slightly contaminated mass spectrum and by mass fragmentography. In addition, 2-MeOEj and OE2-17a were detected in the unconjugated fraction by mass fragmentography, only. The mass spectra obtained for the derivative of OE2 is shown in Fig. 1, and in Fig. 2 the mass spectrum of the trimethylsilyl ether (TMS) derivative of 15a-OHOE2 is compared to that of the correspond¬ ing reference standard. 16,17-epiOE3-TMS (Fig. 3) has a retention time close to that of 15a-OHOE2-TMS on OV-210 and slight contamination due to this compound could be observed in the mass spectrum obtained for 15a-OHOE->TMS (Fig. 2). OE3, OEi and 15a-OHOE2 in the conjugated fraction were identified on the basis of mass spectra which were identical with those of the corresponding reference standards. In addition, the following oestrogens in the conjugated fraction were detected by mass fragmentography: 2-MeOEi, OE2-17/5, 16epiOE3, 16a-OHOE,, 16/Î-OHOE,, 16-oxoOE2 and 15a-OHOE!. In Table 1 the concentrations of the 12 oestrogens identified and quantitated in both the unconjugated and conjugated fractions of the two pools of preg¬ nancy faeces are given. In Table 2, the mean total daily faecal excretion of
The
following oestrogens
) 00
M
1227
-
1358
BO
60 10 i ri
20
160
I.
J
1S6
I| 240
211
Jk
•ft^i"., 100 M/E
ID-CODE
.
€1
051171
MOLECULE WEIGHT t 358. D 1. 0 7. PEAK THRESHOLD i ACC. VOLTAGE . 3.5 KV ELECTRON ENERGIES , 22. 5 EV (11) 2D. 0 EV (1) -
DEPT.
COLUMN
:
SE-30
TEMPERATURES
i
FLASH HEATER 230 C COLUMN 215 C SEPARATOR 250 C -
REL.
OF CLINICAL CHEMISTRY.
Fig.
RET.
-
TIME
:
.59
( CHOLESTANE -
UNIVERSITY OF HELSINKI
1.
Mass spectrum of the methylated and TMS derivative of oestradiol isolated from the unconjugated fraction of human pregnancy faeces.
100
M
;504 BO -
60 -
10
i
-
20 -
,
298 324
Jlr'.^rr-i't'ifAri.lfki M,',
II J4D 345_
270
i
(¡,4 I ,1. I\'>
r>r
^O" ,
X
j
>i
489
-r
r-r4-,
M/E
218 ID-CODE i 021071 MOLECULE WEIGHT 5 01. 0 PEAK THRESHOLD i 1.07. ACC. VOLTAGE : 3. 5 KV ELECTRON ENERGIES ( 20. 0 EV (I) 22. 5 EV (¡I)
COLUMN
DEPT.
i
OV-210
TEMPERATURES
-
FLASH HEATER 230 C COLUMN 220 C SEPARATOR 210 C 1.20 (CHOLESTANE TIME < .
-
-
PEL.
RET.
-
UNIVERSITY OF HELSINKI
OF CLINICAL CHEMISTRY.
100
M
504
80
.-rriU.iiy.,.,.L mwÍ^.U V
399
*
386, 414
lr\i i M/E
120 ID-CODE 010675 MOLECULE WEIGHT .501. 0 PEAK THRESHOLD i 1. 0 7.
COLUMN
ELECTRON ENERGIES = 20. 0 EV (I) 22. 5 EV
REL.
-
ACC.
VOLTAGE
DEPT.
3. 5 KV
:
OV-210
TEMPERATURES
FLASH HEATER 218 C COLUMN 208 C SEPARATOR 216 C TIME 1.23 ( CHOLESTANE .
-
-
til)
OF CLINICAL CHEMISTRY,
Pig.
RET.
-
UNIVERSITY OF HELSINKI
2.
Mass spectrum of the TMS derivative of 15a-hydroxyoestradiol fraction of human pregnancy faeces (top panel) and the corresponding reference standard (lower panel). The compound from faeces is slightly contaminated with 16,17-epioestriol, the mass spectrum of which is seen in Fig. 3.
11 of the
oestrogens is compared
to their
plasma, biliary and urinary concentrations in the last trimester of pregnancy (Adlercreutz et al. \974a,b). The total oestrogen excretion measured was 786 fig/24 h in the day 1 pool and 1300 /,g/24 h in the day 2 pool. Of this 97.8 and 98.6%, respectively, were present in the unconjugated fraction. The only oestrogen preferentially mean
M
100
504 BO 310
60
345
386
I
324
414
360
I.J...U II jJaMX M/E
ID-CODE . 010875 85 MOLECULE WEIGHT .501.0 PEAK THRESHOLD 2. 0 X ACC. VOLTAGE : 3. 5 KV ELECTRON ENERGIES . 20. 0 EV (I) 22. 5 EV (II)
COLUMN
DEPT.
;
OV-210
FLASH HEATER 21B C COLUMN 208 C SEPARATOR 218 C TIME i 1.31 (CHOLESTANE
TEMPERATURES
-
i
-
REL.
OF CLINICAL CHEMISTRY.
RET.
-
1
-
UNIVERSITY OF HELSINKI
Pig.
3.
Mass spectrum of the TMS derivative of
16,17-epioestriol reference
standard.
Table 1.
Oestrogen
concentrations in the
unconjugated
pools of human
and conjugated fractions of two 24-h pregnancy faeces.
Unconjugated Day
Conjugated
Day
1
Day
OE,
2-MeOE, OE2-17/? OE2-17a 16-epiOE3 17-epiOE3 16a-OHOE, 16^-OHOE,
160.5
244.5
0.4
3.6 90.3 22.9 2.8
6.0 194.0 53.6 3.0
