Bo Holmstedt

Mass fragmentography. Identification of chlorpromazine and its metabolites in human blood by a new method.

Abstract Chlorpromazine and some of its metabolites have been identified in human blood with the combination of gas chromatography and mass spectrometry. A new method, mass fragmentography has been elaborated. It is based upon a continuous recording of up to three mass numbers characteristic of a single substance or a group of compounds. With this technique both a high sensitivity and a high selectivity which can be changed according to wish are achieved. Compounds are identified by their retention times and the fact that all mass numbers are represented in characteristic relative intensities. Refocusing on other characteristic fragments and/or the molecular ion confirms the identity. Through repeated refocusing “a partial mass spectrum” of a compound can be established even when the amounts present are too small for the scanning of a complete spectrum. With this method chlorpromazine, and its desmethylated and didesmethylated metabolites have been identified in plasma . The latter two metabolites were obtained also after treatment of the plasma with β-glucuronidase, as was 2-chlorophenothiazinylpropionic acid, which was found in quantities that allowed the scanning of a complete mass spectrum. In red blood cells the two desmethylated metabolites could be identified with mass fragmentography only after treatment with β-glucuronidase. The use of the method is discussed, particularly with regard to blood levels of drugs as related to therapeutic effects and side effects.

Transformation and action of metrifonate

Chemical formation of dichlorvos (2,2-dichlorovinyl dimethyl phosphate) was found to occur in mouse brain after i.p. injection of metrifonate (2,2,2-trichloro-1-hydroxyethyl dimethyl phosphonate). A mass fragmentographic technique was used. Different isotopic variants were used both as internal standards and to compensate for dichlorvos formed during the workup procedure. The dichlorvos formed in vivo was found to have its maximal concentration a few minutes after the maximum of the metrifonate itself.The effect of metrifonate and dichlorvos on acetylcholine levels, acetylcholinesterase activity and synthesis rate of acetylcholine in mouse brain was also studied. In all three cases the effect of metrifonate was found to be prolonged and delayed as compared to the effect of dichlorvos.It is concluded that metrifonate acts as a slow release formulation in the body giving rise to dichlorvos nonenzymatically. This circumstance at least partly explains its efficacy in the treatment of schistosomiasis.

Quantitation of Δ1‐tetrahydrocannabinol in plasma from cannabis smokers

A method to identify and accurately measure non‐labelled Δ1‐tetrahydrocannabinol (Δ1‐THC) in blood of cannabis smokers has been developed. It consists of the following steps: To a 5 ml plasma sample is added deuterated Δ1‐THC (Δ1‐THC‐d2) as internal standard. After extraction with light petroleum and evaporation, the Δ1‐THC containing fraction is separated by chromatography on Sephadex LH‐20 (1 times 40 cm) using light petroleum‐chloroform‐ethanol (10:10:1) as eluant. A fraction containing Δ1‐THC is collected and subjected to mass fragmentography (LKB 9000; 3% OV‐17/Gas‐Chrom Q; 230°). The mass spectrometer was adjusted to record the intensities of m/e 299 and 314 of Δ1‐THC and m/e 301 and 316 of Δ1‐THC‐d2. The standard curve was made by plotting peak height m/e 299/m/e 301. Peak levels of 19–26 ng ml−1 were reached within 10 min after smoking a cigarette containing 10 mg Δ1‐THC.

Cocaine in blood of coca chewers.

Coca leaves (Erythroxylum coca Lamarck) and powder (5 - 10 g) were taken orally by human subjects in the same way as South American natives do. The cocaine, as measured by mass fragmentography, was immediately detected in the blood, reached peak concentrations from 10 - 150 ng/ml plasma at 0.38 - 1.95 hours, and persisted in the plasma for more than 7 hours. Half-lives of the elimination of cocaine were calculated and ranged from 1.0 to 1.9 hours. The absorption half-lives ranged from 0.2 to 0.6 hours. The shape of the curves fits with the subjective effects reported. There is no reason to believe that the stimulating effect achieved by the use of either coca leaves or powder is not due to cocaine.

Gas chromatography‐mass spectrometry of nortriptyline in body fluids of man

The application of the mass fragmentographic technique has aUowed accurate identification of a tricyclic antidepressant drug, nortriptyline, and some of its metabolites in biological fluids from patients treated with therapeutic doses and from one patient intoxicated with the same drug. Mass fragmentography in combination with the scanning of mass spectra of compoundy eluted from the gas chromatography column made it possible to identify nortriptyline and the metabolites, desmethylnortriptyline, 10‐hydroxynortriptyline, and, tentatively, 10‐hydroxydesmethylnortriptyline.

Quantitative Determination of Nortriptyline And Desmethylnortriptyline In Human Plasma By Combined Gas Ciiromatography - Mass Spectrometry

Abstract A quantitative method has been developed for the determination of the tricyclic antidepressant drug nortriptyline in plasma, utilizing a combined technique of gas chromatography - mass spectrometry called mass fragmentography. A metabolite, desmethylnortriptyline, may be analyzed in the same run. The compounds are first converted to their heptafluorobutyranide derivatives. A complete chromatogram takes less than five minutes. Quantitive determinations under the present conditions are possible down to a few nanograms of nortriptyline per milliliter. There is a good correlation between the new method and that previously used which was based upon in vitro 3 11-acetylation labelling.

Determination of cocaine in some south american species of Erythroxylum using mass fragmentography

Abstract Thirteen South American species of Erythroxylum have been analyzed for their cocaine content. Cocaine was found only in E. coca Lam., E. novogranatense (Morris) Hieron. and E. novogranatense var. truxillense (Rusby) Machado. The amount of cocaine was determined by mass fragmentography using deuterium labelled cocaine as internal standard

Prolegomena to Seveso

Worldwide attraction was bestowed upon the accident that occurred at the ICMESA factory in the small Italian community of Meda/Seveso at 12.30 on July 10th, 1976. This has resulted in a host of publications, including half a dozen books, at the time of writing. The pre-history of this event is also discussed in many publications, as is the use of the herbicide 2,4,5-T in agriculture and as a defoliant in the Vietnam war. This present account will not attempt to go into detail except for some aspects of the protohistory of Seveso and the chronology of the events occurring immediately after the explosion at the ICMESA factory. The author has interviewed several people and consulted a number of publications in an effort to try and throw some light on facts such as why the publication of the formula of one of the most toxic compounds ever described and published in 1957, attracted so very little attention for so many years, and why the more than 20 cases of intoxication with TCDD, which had occurred prior to the Seveso (Italy) disaster, had remained relatively unknown in spite of information found in open literature. In the case of ICMESA, representatives have been interviewed from both the research workers of the chemical companies concerned and Italian scientists dealing with the problems of chemistry and toxicology occasioned by the Seveso accident. The author has also taken information from a report made by an independent Swedish commission sent to Seveso (Italy) shortly after the accident (Karnedo report). This commission was triggered by an organization that has the ability to send experienced people to investigate, without delay, any reported medical catastrophy in the world. Many letters sent to the author will also be quoted in this account. Because of the nature of this paper relatively few references will be found in the text. Instead, the list of references has been partly annotated. The present report does not discuss the problem of dioxins as contaminants in defoliants used in the Vietnam war. In this case the reader is referred to the publications of

MOBILIZATION OF ACETYLCHOLINE BY CHOLINERGIC AGENTS

Many compounds are known to interfere with acetylcholine (AcCh) metabolism, but relatively few experiments have been reported where AcCh has been measured quantitatively in the brain, and where the values have been related to the symptomatology. Three groups of compounds will be treated here, namely: a ) cholinesterase inhibitors b ) barbiturates c ) tremorgenic agents (oxotremorine and arecoline) They differ in that they are differently influenced by atropine when the latter agent is given before or after the respective compounds.

TREMORGENIC AGENTS AND BRAIN ACETYLCHOLINE

Publisher Summary This chapter discusses the tremorgenic agents and brain acetylcholine. The most important role of the tremorgenic agents can well be the light they may be able to throw upon transmission in the central nervous system. Acetylcholine was the only brain amine which in experience could be appreciably influenced by the tremorgenic agents during the period of tremor. The time of tremor activity also roughly corresponds to the rise in brain acetylcholine. In all three instances the peak activity of the tremor occurs, however, before that of brain acetylcholine. This is most marked with the long acting compound tremorine that has been proven to be metabolized in the body. It is less evident with oxotremorine and arecoline. On the other hand, the rise in tremor activity is concomitant with the rise in brain acetylcholine. Dose–response curves with regard to increase in brain acetylcholine were established for all three agents. Arecoline was most potent in this respect increasing brain acetylcholine to about 25 μ moles per gram tissue.