Birgitta Sjöquist

Identification of prostaglandin D2 as a major prostaglandin in homogenates of rat brain

Prostaglandin (PG) E2, D2, F2alpha and thromboxane B2 (TxB2) were determined in homogenates of rat brain by gas-chromatography--mass spectrometry. The level of PGD2 was 735 +/- 19 ng/g, of PGF2alpha 150 +/- 13 ng/g, of TxB2 112 ng/g and of PGE2 86 +/- 8 ng/g. The same relative proportions of cyclooxygenase products were found in incubates of unstimulated sliced rat brain. 14C-PGH2 was converted in high yield into PGD2 by enzyme(s) present in the soluble fraction of the homogenate. These results indicate that PGD2 is the major cyclooxygenase product in the central nervous system of the rat.

Increased salsolinol levels in rat striatum and limbic forebrain following chronic ethanol treatment

Abstract: Endogenous levels of salsolinol and its methylated metabolite were measured by combined gas chromatography and mass spectrometry in rats chronically exposed to ethanol for 150 days. The chronic ethanol administration produced a significant increase of salsolinol concentrations in dopamine‐rich brain areas, e.g., the striatum and the limbic forebrain. A negative correlation was observed between plasma ethanol concentration and the level of salsolinol in the brain. A possible role for salsolinol in the regulation of ethanol drinking and/or in the development of ethanol dependence is discussed.

Mass fragmentographic determination of 4-hydroxy-3-methoxyphenylglycol (HMPG) in urine, cerebrospinal fluid, plasma and tissues using a deuterium-labelled internal standard

4-Hydroxy-3-methoxyphenylglycol (HMPG) with the three hydrogen atoms in the side-chain replaced with deuterium (HMPG-D3) was used as the internal standard in the mass fragmentographic determination of free and conjugated HMPG in human urine, cerebrospinal fluid and plasma and in rat urine, liver and brain. HMPG-D3 was added to body fluids or homogenates followed by enzymatic hydrolysis of conjugates. HMPG was extracted with ethyl acetate, converted into the trifluoroacetyl derivative and analyzed by mass fragmentography. HMPG levels were 7.4 nmoles/ml plus or minus 2.8% in human urine, 73 pmoles/ml plus or minus 8.2% in human cerebrospinal fluid, 56 pmoles/ml plus or minus 5.4% in human plasma, 24 nmoles/ml plus or minus 3.6% in rat urine, 0.26 nmoles/g plus or minus 6.2% in rat brain and 99 pmoles/g plus or minus 13% in rat liver. The method is highly specific and sensitive, permitting analysis in small samples or in plasma and in tissues for which previously no methods for HMPG analysis were available.

Analysis of salsolinol and salsoline in biological samples using deuterium-labelled internal standards and gas chromatography—mass spectrometry

Salsolinol and salsoline were labelled with deuterium using an acidic exchange reaction in 2HCl--2H2O. Two deuterium atoms were incorporated in both compounds. The deuterium-labelled salsolinol and salsoline were used as internal standards to determine picomol amounts of the corresponding unlabelled compound in the urine, cerebrospinal fluid, brain and liver. The salsolinol was purified on alumina and salsoline collected in the effluent. The compounds were analysed as their pentafluoropropionyl derivatives by gas chromatography on a 1% OV-17 column and were selectively detected with electron-impact mass spectrometry at the molecular ions M+ and M+--15. With human urine the precision of the methods were +/- 4.9% (coefficient of variation, n = 10) for salsolinol and +/- 2.2% for salsoline at a level of 0.100 nmol/ml. Administration of salsolinol to rats intraperitoneally (0.4 mmol/kg) resulted in levels of 1--2 nmol/g in striatum and limbic forebrain after 2 h, whereas the corresponding liver values were about 550 nmol/g. Control animals showed salsolinol values in liver of about 2 nmol/g and in striatum and limbic forebrain 1 nmol/g tissue.

Norepinephrine Metabolism in Man Using Deuterium Labelling: The Conversion of 4-Hydroxy-3-Methoxyphenylglycol to 4-Hydroxy-3-Methoxymandelic Acid

Abstract: 4‐Hydroxy‐3‐methoxyphenylglycol (HMPG) labelled with three deuterium atoms was used to study the disposition of peripherally administered HMPG. Five healthy men were given an intravenous pulse dose of 4.3 μmol of labelled HMPG and subsequent plasma and urine levels of endogenous and labelled HMPG as well as those of 4‐hydroxy‐3‐methoxymandelic acid (HMMA, VMA) were determined by gas chromatography‐mass spectrometry, using selected ion detection. Approximately 40% of the injected amount of deuterium‐labelled HMPG was recovered in the urine as HMMA and another 40% was eliminated as HMPG conjugates. Thus, the HMPG formed from norepinephrine either in the central or peripheral nervous system undergoes both conjugation and extensive oxidation.

The effect of alcoholism on salsolinol and biogenic amines in human brain

Chronic alcoholics with a well documented history of alcohol abuse were divided at autopsy into two groups, intoxicated alcoholics (IA, with blood ethanol levels) and sober alcoholics (SA, without blood ethanol levels). Norepinephrine (NE), 4-hydroxy-3-methoxyphenyl-glycol (HMPG), 5-hydroxyindoleacetic acid (5-HIAA), dopamine (DA), salsolinol (SAL) and methylated salsolinol (M-SAL) in caudate nucleus and putamen from the two groups of alcoholics and an age-matched control group were analysed with gas chromatography-mass spectrometry. HMPG was significantly increased in the brains of the alcoholics compared to controls (IA, P less than 0.001 and SA, P less than 0.05) indicating an increased NE turnover in the brains of alcoholics. 5-HIAA, the major 5-hydroxytryptophan (5-HT) metabolite, was increased in the caudate nucleus from SA (P less than 0.05) compared to controls and IA. This could be interpreted as an increased turnover of 5-HT at abstinence. SAL can be formed from DA and acetaldehyde and/or pyruvate. SAL was found in all brains studied. The levels were about 1% of the dopamine concentrations. SAL had a tendency to be increased in brains of IA compared to control brains. The most remarkable finding was that SAL was significantly decreased in both the caudate nucleus and putamen from SA compared to controls and IA. M-SAL showed tendencies in the same directions as SAL. DA did not show any significant changes, but tended to be lower in SA.(ABSTRACT TRUNCATED AT 250 WORDS)

Determination of homovanillic acid turnover in man

Abstract Homovanillic acid (HVA) labelled with five deuterium (d) atoms was used to determine the total body turnover of HVA, the size of the peripheral body pool of HVA and HVA elimination characteristics in five healthy men. After i.v. injection of 5.5 μmoles (1 mg) of HVA-d 5 the levels of HVA-d 5 and endogenous HVA (HVA-d o ) in plasma and urine were followed by mass fragmentography using HVA-d 2 as the carrier and internal standard. Following an initial distribution phase of 10–20 minutes the plasma elimination curve of HVA-d 5 was monoexponential with a mean T 1 2 of 0.66 hrs. The apparent volume of distribution (V D ) approximated the volume of the body water. The content of HVA in the peripheral body pool calculated from the plasma levels of HVA-d o and V D was 3.4 moles. The urinary HVA excretion rate (mean 1.70 moles/hour) was 45% of the total body turnover, the recovery of urinary HVA-d 5 was 48% of the mean body clearance. Together the results indicate that about 50% of the HVA formed in the body is eliminated by mechanisms other than renal excretion.

Identification and quantification of 1-carboxysalsolinol and salsolinol in biological samples by gas chromatography-mass spectrometry

1-Carboxysalsolinol was found to be present in rat striatum, human urine and caudate nucleus of post mortem human brain, according to capillary column gas chromatographic retention times and selected ion monitoring of the hexafluoropropionyl ester pentafluoropropyl derivative. Simultaneous quantification of 1-carboxysalsolinol and salsolinol was performed in biological samples using deuterium labelled internal standards. In human urine, the precision of the method was +/- 7.1% (coefficient of variation, n = 25) for 1-carboxysalsolinol at 15 pmol/ml and +/- 8.5% for salsolinol at 10 pmol/ml. According to enzymatic hydrolysis, 68% of 1-carboxysalsolinol was found as conjugates in urine, and the corresponding figure for salsolinol was 92%. In human caudate nucleus, the amounts of 1-carboxysalsolinol were found to be significantly greater in brains from alcoholics, who at autopsy had ethanol present in the blood, whereas alcoholics without blood ethanol levels at autopsy had significantly lower concentrations of salsolinol.

Catecholamine metabolites in amniotic fluid as indicators of intrauterine stress.

The catecholamine metabolites HMPG and VMA have been determined in samples of amniotic fluid in 38 uncomplicated pregnancies, seven cases of IUGR, and six cases of diabetes. A successive increase of HMPG and VMA was found toward the end of the pregnancy. HMPG and particularly the HMPG/VMA ratio were significantly higher in the amniotic fluid of the growth-retarded fetuses than in the uncomplicated cases. No significant difference was found between the diabetic and uncomplicated cases.

Mass fragmentographic determination of homovanillic acid in tissues and body fluids using the deuterium labeled species as internal standard

Abstract Mass fragmentographic methods for determination of 4-hydroxy-3-methoxyphenyl acetic acid (HVA) in human cerebrospinal fluid (CSF), urine, blood plasma and mouse brain were developed. After isolation by extraction or by Amberlite XAD-2 chromatography the HVA was converted to the heptafluorobutyryl methyl ester derivative and analyzed using the 2, 2 dideutero-2 (4-hydroxy-3-methoxy-2, 5, 6-trideuterophenyl) acetic acid (HVA-d 5 ) as an internal standard. The values (mean ± coefficient of variation) obtained on repetitive analyses of the same sample were for human CSF 0.42 nmole/ml ± 1.5% (n = 10), human plasma 48 pmole/ml ± 4.5% (n = 15), human urine 20 nmole/ml ± 5.4% (n = 10) and mouse brain 1.2 nmole/g ± 0.6% (n = 12). The results demonstrate that the use of HVA-d 5 as an internal standard provides high precision and the necessary sensitivity for the mass fragmentographic determination of HVA in small amount of tissue and body fluids.