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

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-d₅) 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-d₅ 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.     

The effect of probenecid on catecholamine metabolites in human cerebrospinal fluid analyzed by mass fragmentography.

A gas chromatography-mass fragmentography (GC-MS) method was used to measure homovanillic acid (HVA), vanillylmandelic acid (VMA) and 3-methoxy-4-hydroxyphenethylene glycol (MHPG) in lumbar cerebrospinal fluid (CSF) of 31 patients before and after treatment with probenecid. HVA values increased from 24.6 ± 2.6 S.E.M. to 210 ± 17 ng/ml. The increase in VMA was from 1.06 ± 0.23 to 2.22 ± 0.17 ng/ml and that of MHPG was from 12.2 ± 1.08 to 15.6 ± 1.27 ng/ml. All increases were significant (p = < .01). The results for MHPG and HVA are consistent with results of earlier studies using different methods. VMA concentrations increased significantly but at a rate much lower than those of HVA.     

Conjugated 3,4 dihydroxy phenyl acetic acid (DOPAC) in human and monkey cerebrospinal fluid and rat brain and the effects of probenecid treatment

Gas chromatography-mass spectrometry (GC-MS) was used to measure 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in cerebrospinal fluid from humans and monkeys and in rat caudate nuclei. DOPAC was found to be present mainly in conjugated form. In human lumbar CSF the average concentration of total DOPAC before probenecid treatment was 1.48 ± 0.31 ng/ml; after probenecid it increased to 15.06 ± 3.17 ng/ml. This increase was mainly due to conjugated DOPAC but increases in free DOPAC also occurred. There is a relatively greater accumulation of DOPAC than of HVA, suggesting that in human CSF conjugated DOPAC may have a faster turnover rate than HVA. In monkey, ventricular CSF contained higher concentrations of DOPAC and HVA than did lumbar CSF.In rat brain, treatment with probenecid caused increases in DOPAC, HVA and their conjugates.These results suggest that DOPAC is conjugated in brain and that both compounds are removed from brain and CSF by a probenecid-sensitive acid transport system in the same manner as is HVA.     

Urinary homovanillic acid (HVA) and vanillymandelic acid (VMA) in workers exposed to manganese dust

The neurotoxicity of manganese (Mn) is well known, however, the neurochemical effect caused by this metal is less well investigated. In this study, urinary homovanillic acid (HVA) and vanillymandelic acid (VMA), two end products of catecholamine metabolism, were measured in 39 workers chronically exposed to Mn in a manganese smelting plant. The average duration of Mn exposure was 17.4 yr. Nineteen nonexposed workers were also studied. Concentrations of Mn in serum (MnS) and in urine (MnU) were measured by Zeeman graphite furnace atomic absorption spectrophotometry (ZAAS), and HVA and VMA determined by high performance liquid chromatography (HPLC). For Mn-exposed workers, the concentration of MnS was nearly 2.8 times (1.61 ± 0.16 mg/L vs 0.56 ± 0.16 mg/L) and MnU about 4.5 times higher (7.62 ± 0.17 mg/L vs 1.69 ± 0.16 mg/L) than the nonexposed. Although the geometric mean concentration of HVA in exposed workers was similar to that of the nonexposed (3.09 ± 1.39 mg/g ere. vs 2.99 ± 1.40 mg/g cre.), the VMA concentration was significantly higher (3.02 ± 1.43 mg/g cre. vs 2.49 ± 1.58 mg/g cre.,p = 0.033). Multiple regression analysis showed that although there were no correlations between any of these parameters with the duration of exposure to Mn, both HVA and VMA showed significant correlations with increase in MnS and MnU. These data provide evidence that exposure to Mn was associated with measurable increase in catecholamine metabolites. This finding is compatible with recent observations in laboratory animals that Mn interferes with neurochemical metabolism.     

Dopamine and Serotonin Metabolism in Hepatic Encephalopathy

Patients with stupor or coma from fulminant hepatic failure were found to have high cerebrospinal fluid concentrations of homovanillic acid (HVA) and 5-hydroxyindole acetic acid (5-HIAA), metabolites of dopamine and serotonin respectively. Excessive amounts of their precursors—phenylalanine and tyrosine and free tryptophan—were found in the patients'' plasma. Methionine, which participates in dopamine degradation, was also much increased. Similar disturbances were found in patients suffering an acute exacerbation of chronic encephalopathy. These abnormalities would be consistent with other evidence of an increased turnover of serotonin and possibly dopamine in the brain during hepatic encephalopathy.     

Influence of dopamine agonists on plasma and brain levels of homovanillic acid

The response of the plasma dopamine (DA) metabolite, homovanillic acid (HVA), to two DA agonists was investigated in the rat. Apomorphine administered i.p. (2 mg/kg) produced, within one hour, a significant decrease in plasma HVA. The response of plasma HVA to apomorphine was also investigated after pretreatment with debrisoquin, a drug which selectively blocks peripheral HVA production by inhibition of MAO. Pretreatment with debrisoquin did not significantly alter the decrement in plasma HVA produced by apomorphine indicating that a substantial portion of the plasma HVA response to apomorphine is due to the drug's action on brain. Bromocriptine (2 mg/kg) was also found to produce a significant decrease in plasma HVA. Since the response of brain HVA to DA agonists reflects the sensitivity of the DA receptor, the plasma HVA response to DA agonists might be a practical method of assessing brain DA receptor sensitivity in humans.     

Mass fragmentographic assay of homovanillic acid in brain tissue

Abstract— Available methods for the determination of homovanillic acid (HVA) are based on fluorimetric measurements. The present method uses gas chromatography-mass spectrometry with the aid of deuterium labelled HVA methyl ester as a carrier and internal standard. Quantitation is achieved by comparing intensities of the molecular ion (= base peak) of the protium (H) and deuterium (D) forms of the methyl ester heptafluorobutyric derivatives of HVA. A standard curve is constructed by measuring mixtures of known amounts of protium and deuterium derivatives and plotting peak height ratios H/D on the ordinate and ratios of amounts H/D on the abscissa. Analysis of ten individual mouse brains gave a mean value of 1·36 nmole/g brain tissue with a standard deviation of ±9 per cent. Chlorpromazine elevates, whereas pargyline reduces the level of HVA in brain markedly. The described procedure offers advantages because of the greatly increased specificity and sensitivity permitting analyses in discrete brain areas.     

Determination of the Lumped Constant for the α-Methyltryptophan Method of Estimating the Rate of Serotonin Synthesis

Abstract: The lumped constant (LC) for the α-methyl-l -tryptophan method to convert the brain's uptake of labeled α-methyl-l -tryptophan into the regional rate of serotonin synthesis was estimated. The method involved independently estimating the unidirectional uptake constant of the tracer (α-[¹⁴C]methyl-l -tryptophan) to the tissue and the tracee (tryptophan) (with the addition of a radioactive compound) and calculating their ratio. The LC was estimated from logarithmically transformed data. Similar experiments were performed using rats treated with the drug probenecid, which blocks the efflux of 5-hydroxyindoleacetic acid (a metabolite of serotonin) from the brain. The experiments using probenecid, corrected for the difference in the levels of plasma free tryptophan (increased in probenecid-treated rats) relative to control experiments, gave an average LC for the rat brain of 0.46 ± 0.14 (mean ± SD). This value was not significantly different from the one obtained in controls (0.43 ± 0.13). In addition, the LC was also calculated using unidirectional uptake constants in the probenecid-treated rats for α-methyl-l -tryptophan and l -tryptophan. This LC value was 0.39 ± 0.10. There was no significant difference between these three LC values. Thus, an average ± SD LC of 0.42 ± 0.07 for 28 brain structures investigated in this study was obtained. Statistically the LC obtained in different structures had a variability that could be accounted for by errors in measurements alone. In other words, dispersion in the LC values could be fully accounted for by chance alone. Data confirmed that the LC value did not change when the rate of serotonin synthesis was increased by probenecid treatment. We also showed that the rate of 5-hydroxyindoleacetic acid accumulation in probenecid-treated rats was 58 pmol g^?1 min^?1 (rat brain), which is about twice as much as reported by others for a normal rat. This difference could also be accounted for by the increase in the plasma level of free tryptophan in probenecid-treated rats.     

Kinetics of Homovanillie Acid and Determination of Its Production Rate in Humans

Abstract: Homovanillie acid (HVA) labeled with either two or five deuterium atoms (d₂-HVA or d₅-HVA) was used to label the peripheral body pool of endogenous HVA (d₀-HVA) in control humans and in neurological patients. Either d₂- or d₅-HVA was rapidly injected intravenously and concentrations of d₂- or d₅- and d₀-HVA in sequential samples of blood plasma were determined by gas chromatography-mass spectrometry (GC-MS). Parameters describing the distribution and elimination of HVA, as well as its pool size, turnover, and synthesis rate were then calculated. Results indicate that the decline of plasma d₂- or d₅-HVA with time was multiexponential in five out of six subjects. Basal levels of endogenous HVA averaged 14.4 ± 1.3 ng/ml in the control patients and 8.69 ± 2.4 ng/ml in the neurological patients. The biological half-life averaged 71.3 ± 8.0 min in the control subjects and 91.3 ± 15 min in the neurological patients. The apparent volume of distribution of HVA in the body was 31–100 liters. Plasma clearance was 243–679 ml/min. The size of the peripheral body pool, calculated from plasma kinetic parameters, was 392–673 μg. The HVA rate of production, calculated for the whole body, was 166–323 μg/h. This technique can be used to determine accurately the rate of HVA production by the whole body over short time intervals. Since sample size was very limited ( n = 3 per group) and effects of variables such as age and sex on these data have not been excluded, more thorough investigations are needed to document any differences between normal controls and neurological patients.     

A new approach to biochemical evaluation of brain dopamine metabolism

1. Dopaminergic neurotransmission in brain is receiving increased attention because of its known involvement in Parkinson's disease and new methods for the treatment of this disorder and because of hypotheses relating several psychiatric disorders to abnormalities in brain dopaminergic systems. 2. Chemical assessment of brain dopamine metabolism has been attempted by measuring levels of its major metabolite, homovanillic acid (HVA), in cerebrospinal fluid, plasma, or urine. Because HVA is derived in part from dopamine formed in noradrenergic neurons, plasma levels and urinary excretion rates of HVA do not adequately reflect solely metabolism of brain dopamine. 3. Using debrisoquin, the peripheral contributions of HVA to plasma or urinary HVA can be diminished, but the extent of residual HVA formation in noradrenergic neurons is unknown. By measuring the levels of methoxy-hydroxyphenylglycol (MHPG) in plasma or of urinary norepinephrine metabolites (total MHPG in monkeys; the sum of total MHPG and vanillyl mandelic acid (VMA) in humans) along with HVA, it is possible to estimate the degree of impairment by debrisoquin of HVA formation from noradrenergic neuronal dopamine and thereby better assess brain dopamine metabolism. 4. This method was applied to a monkey before and after destruction of the nigrostriatal pathway by the administration of MPTP.     

TURNOVER OF FREE AND CONJUGATED (SULPHONYLOXY) DIHYDROXYPHENYLACETIC ACID AND HOMOVANILLIC ACID IN RAT STRIATUM

Abstract— Conjugated (sulphonyloxy) dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) were synthesized from free DOPAC and HVA and used as reference compounds in their fluorimetric determination in rat brain (detection limit 0.2 nmol/g). The conjugated DOPAC and HVA form 29 and 36% of the total DOPAC and HVA found in rat striatum, respectively. Dopamine (DA) metabolism was studied in the rat striatum by following the decline of both free and conjugated DOPAC and HVA after treatment with pargyline (100mg/kg. i.p.) either alone or in combination with tropolone (100 mg/kg, i.p.). or from the accumulation of the free and conjugated acids after treatment with probenecid (100-500mg/kg. i.p.). The rates of decline were analysed by a non-linear curve fitting method using a simple model of DA metabolism that postulates the formation of the conjugates exclusively from the free acids, and HVA from DOPAC, with first order kinetics and single open compartments only. The curves computed all passed through the s.e.m. of every experimental point. The rate constants thus found indicate that DOPAC turnover is about 23nmol/g/h. Of this about 16 nmol/g/h are O -methylated to HVA, about 6 nmol/g/h are conjugated and less than 1 nmol/g/h is eliminated as free DOPAC. Of the HVA formed, about 8.5nmol/g/h are conjugated and about 7.5 nmol/g/h eliminated as free HVA. The conjugates accumulated after treatment with probenecid (1 h) faster than the free acids. The maximal accumulation of all four metabolites found (21 nmol/g/h) approximates the total turnover of DOPAC.     

Effect of a test meal,duodenal acidification,and tetragastrin on the plasma concentration of β-endorphin-like immunoreactivity in man

The effects of various test materials on plasma β-endorphin-like immunoreactivity (β-EpLI) were investigated in man using a specific radioimmunoassay developed by the authors. Plasma β-EpLI was determined after extraction by the acid/acetone method (recovery 73±5%). The intraassay and interassay coefficients of variation were 5.0% and 7.6%, respectively. The plasma concentrations of human β-EpLI in normal subjects were 11.6±4.0 pmol/l for men (n=23) and 10.7±4.8 pmol/l for women (n=27). Ingestion of a test meal (150 g of Campbell's condensed meat soup) resulted in a biphasic rise in plasma β-EpLI from the basal level of 4.4±1.0 pmol/l to 29.2±1.9 pmol/l after 5 min and 24.8±6.7 pmol/l after 90 min. Intraduodenal infusion of 115 ml of 0.1 M HCl over 10 min increased the plasma β-EpLI level from 8.7±0.5 pmol/l to 15.5±0.4 pmol/l at 10 min after the start of infusion, but the level rapidly returned to the initial value after the end of the infusion. Intramuscular injection of 4 μg/kg body weight of tetragastrin markedly stimulated gastric acid output and β-EpLI release, but pretreatment with 10 mg of histamine H₂ receptor antagonist inhibited the gastric acid output and plasma β-EpLI release induced by tetragastrin.These results indicate that β-EpLI release is stimulated by ingestion of meat soup, duodenal acidification and tetragastrin administration. It is suggested that gastric acid participates, at least in part, in postprandial release of β-EpLI, probably from the gastrointestinal tract.     

Determination of homovanillic acid turnover in man

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-d₅ the levels of HVA-d₅ and endogenous HVA (HVA-d_o) in plasma and urine were followed by mass fragmentography using HVA-d₂ as the carrier and internal standard. Following an initial distribution phase of 10–20 minutes the plasma elimination curve of HVA-d₅ was monoexponential with a mean $\text{T}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$ 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-d₅ 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.     

Relationships of Dopamine, Cortical Oxygen Pressure, and Hydroxyl Radicals in Brain of Newborn Piglets During Hypoxia and Posthypoxic Recovery

Abstract: The present study describes the relationships of extracellular striatal dopamine, cortical oxygen pressure, and striatal hydroxyl radicals in brain of newborn piglets during hypoxia and posthypoxic reoxygenation. Hypoxia was induced by reducing the fraction of inspired oxygen (F_iO₂) from 22% (control) to 7% for 1 h. The F_iO₂ was then returned to the control value and measurements were continued for 2 h. Cerebral oxygen pressure was measured by the oxygen dependent quenching of phosphorescence and extracellular levels of dopamine, 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), and hydroxy radicals in the striatum were determined by in vivo microdialysis. Hypoxia decreased the cortical oxygen pressure from 47 ± 2 to 9 ± 1.3 torr (p < 0.001); the levels of extracellular dopamine in the striatum increased to 16,000 ± 3,270% of control (p < 0.01), whereas the levels of DOPAC and HVA decreased to 25.3 ± 6% (p < 0.001) and 36 ± 5% (p < 0.01) of control, respectively. Compared with control, the hydroxyl radical levels at each time point were not significantly increased during hypoxia, although the sum of the measured values was significantly increased (p < 0.05). During the first 5 min after F_iO₂ was returned to 22%, the cortical oxygen pressure increased to control values and stayed at this level for the remainder of the measurement period. The extracellular level of dopamine declined to values not statistically different from control during 40 min of reoxygenation. During the first 10 min of reoxygenation, DOPAC and HVA further decreased and then began to slowly increase. By 70 min of reoxygenation, the values were not significantly different from control. Hydroxyl radicals were above control during the entire period of reoxygenation, with maximal values observed after 100 min of reoxygenation. This increase was largely abolished by injecting the animals with α-methyl-p-tyrosine 5 h before hypoxia, a procedure that depleted the brain of dopamine. Our results suggest that oxidation of striatal dopamine during posthypoxic reoxygenation is at least partly responsible for the observed increase in striatal level of hydroxyl radicals that may exacerbate posthypoxic cerebral injury.     

Antiserum to 5alpha-dihydrotestosterone: production, characterization and use in radioimmunoassay

5α-Dihydrotestosterone (5α-DHT) was rendered antigenic by covalent attachment to bovine serum albumin (BSA) through position 1 of the steroid. Nucleophilic attack by β-mercaptopropionic acid on the 1,2-dehydro derivative of 5α-DHT yielded the corresponding 1α-thioether alkanoic acid which was coupled to bovine serum albumin by use of the carbodiimide reagent. The method should be generally applicable to 3-oxosteroids. Immunization of rabbits with 5α-DHT-1α-carboxyethyl-thioether-BSA gave rise to antisera of high affinity for 5α-DHT (K_a= 1.4 × 10⁹ 1/mol) that showed little cross reaction with 17β-hydroxy-5β-androstan-3-one (3%), and with a variety of 17-oxoandrostane compounds (≤0.5%). However the serum cross-reacted significantly with testosterone (10%) and with 5α-androstene-3α, 17β-diol (16%). A radioimmunoassay procedure for the determination of 5α-DHT in plasma is described. Chromatographic purification of the plasma extracts proved necessary for obtaining valid results. The plasma level of 5α-DHT(pg/ml; ean ± S.D.) was 364±79 (n = 7) in normal human adult males and 188 ± 62 (n = 5) in normal non-pregnant women.     

A DIRECT METHOD FOR DETERMINING DOPAMINE SYNTHESIS AND OUTPUT OF DOPAMINE METABOLITES FROM BRAIN IN AWAKE ANIMALS

Abstract— A direct method for measuring the rate of dopamine (DA) synthesis and the DA metabolites by the brain of awake monkeys ( Macaca arctoides ) is described. The method utilizes a coupling of a measure of cerebral blood flow with the mass spectrometrically determined difference in the concentrations of the metabolite under study in plasma obtained from arterial and internal jugular bulb blood. For homovanillic acid (HVA) a consistent and highly significant veno-arterial (V-A) difference of 2.2 ± 0.4 ng/ml of plasma ( P < 0.0005) was found. When this V-A difference was coupled with a measure of cerebral blood flow it was determined that, in the awake monkey, the average output of HVA by brain was 113.4 ± 19.1ng/100g brain min⁻¹. There were large individual variations, however, between animals (range = 38-194 ng/100g brain min⁻¹). In contrast to HVA, no consistent V-A difference for dihydroxyphenylacetic acid (DOPAC) was found; i.e. the concentrations of DOPAC in plasma obtained from arterial and internal jugular bulb venous blood were essentially identical. These data indicate that, in contrast to the rat, in this non-human primate HVA is the major metabolic product of brain DA. Since HVA is the major metabolite of DA, production of HVA under steady state conditions gives a measure of DA synthesis by whole brain; i.e. the rate of DA synthesis by whole brain in the awake monkey is 113.4 ± 19.1ng/100g brain min⁻¹. It is suggested that this technique may be of value in both basic and applied types of studies.     

Plasma homovanillic acid as an index of brain dopamine metabolism: Enhancement with debrisoquin

Plasma levels of the dopamine (DA) metabolite homovanillic acid (HVA) may be a useful measure of brain HVA production by central DA systems. Even though there is a significant peripheral contribution to plasma HVA, experimental manipulations that alter brain HVA produce parallel changes in plasma HVA levels. This study was designed to assess whether the ability of plasma HVA to reflect haloperidol induced increases in brain HVA could be strengthened by reducing the contribution to plasma HVA from peripheral sources. Debrisoquin sulfate, a monoamine oxidase inhibitor that does not enter the brain, was given in a low dose schedule to rats and lowered the peripheral contribution to plasma HVA by between 42 and 68%, resulting in a situation where between 62 and 87% of plasma HVA derived from brain. Using this dose schedule, rats pretreated with debrisoquin displayed a significant increase in plasma HVA following a lower dose of haloperidol than that required in the vehicle pretreated rats. In the debrisoquin pretreated group, a 71% increase in brain HVA was accompanied by a significant 60% increase in plasma HVA, whereas the vehicle pretreated group required a 136% increase in brain HVA to display a significant 50% increase in plasma. These findings indicate that debrisoquin pretreatment improves the reliability of plasma HVA to reflect changes in brain DA metabolism. Plasma HVA samples obtained from humans following debrisoquin may provide a clinically applicable method for assessing brain DA systems in neurologic and psychiatric illness.     

Effects of an N-methyl-d-aspartate receptor agonist and its antagonist CPP on the levels of dopamine and serotonin metabolites in rat striatum collected in vivo by using a brain dialysis technique

3-((±)-2-Carboxypiperazin-4-yl)propyl-1-phosphonic acid (CPP) is an antagonist at the N-methyl-D-aspartate (NMDA) subtype of glutamate receptor. In the present study, levels of dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA) and 5-hydroxyindolacetic acid (5-HIAA) were measured after intracerebroventricular injection of NMDA, CPP or both in rat striatum using a brain dialysis method. The injection of NMDA produced a significant increase in DOPAC level. HVA level was also increased by NMDA injection. The level of 5-HIAA was not affected by NMDA injection. The injection of CPP had no effect on DOPAC, HVA and 5-HIAA levels. The injection of CPP restrained the increase of DOPAC and HVA levels induced by NMDA injection. The results suggest that intracerebral injection of NMDA may increase dopamine release from rat striatum, but have no effect on serotonin release. Furthermore, CPP inhibits NMDA induced release of dopamine.     

CORRELATIONS BETWEEN A FLUORIMETRIC and MASS FRAGMENTOGRAPHIC METHOD FOR THE DETERMINATION OF 3-METHOXY-4- HYDROXYPHENYLACETIC ACID and TWO MASS FRAGMENTOGRAPHIC METHODS FOR THE DETERMINATION OF 3-METHOXY-4- HYDROXYPHENYLETHYLENE GLYCOL IN CEREBROSPINAL FLUID

One-hundred and twenty-two lumbar cerebrospinal fluid (CSF) samples were assayed for 3-methoxy-4-hydroxyphenylacetic acid (HVA) by both a fluorimetric and mass fragmentographic method. The correlation coefficient (cc) and residual standard deviation (Syx) of the results were calculated as 0.966 and 23.3 ng/ml, respectively. If only samples containing less than 100ng/ml of HVA were considered, somewhat different values for cc and Syx were found (0.854 and 10.0 ng/ml, respectively). The data obtained by the fluorimetric method were consistently 17% lower than those obtained by the mass fragmentographic method. Spiking experiments resulted in 96.5 ± 7.8% recovery for the fluorimetric method, whereas the use of a deuterated internal standard was found to compensate completely for losses in the mass fragmentographic method. In addition the correlation between two different mass fragmentographic methods for the simultaneous determination of HVA and 3-methoxy-4-hyd-roxyphenylethylene glycol (MOPEG) in CSF was studied. The measurements were performed in different laboratories and resulted in correlation coefficients of 0.941 and 0.940 and residual standard deviations of 7.6 and 1.0 ng/ml for HVA and MOPEG, respectively. From all data we conclude that mass fragmentographic methods for the determination of catecholamine metabolites in CSF are superior to fluorimetric methods because of their selectivity, reproducibility and accuracy.     

Comparative analysis of indices of central dopaminergic functions in man

Various postulated indices of central dopaminergic activity - cerebrospinal fluid (CSF) dopamine (DA), dihydroxy-phenylacetic acid (DOPAC), homovanillic acid (HVA), noradrenaline (NA), plasma NA, serum prolactin, serum dopamine-β-hydroxylase (DBH), and platelet monoamine oxidase (MAO) activity - were measured in 30 drug-free inpatients. The mean values and the ranges were similar to those described in the literature. Plasma NA showed significant positive correlation with age. Significant positive correlation was found between CSF DA and its metabolites DOPAC and HVA. Serum DBH activity showed a slight but significant inverse correlation with CSF DA and its two metabolites. CSF NA showed a significant positive correlation with CSF DOPAC, but only in females. Serum DBH activity had no significant correlation either with CSF or with plasma NA levels. These findings suggest that either CSF HVA or DOPAC and DA may be useful indicators of DA metabolism in humans. Serum DBH activity may be in relationship with the central dopaminergic functions.