Review
Metabolism and metabolomics of opiates: A long way of forensic implications to unravel

https://doi.org/10.1016/j.jflm.2018.12.005Get rights and content

Highlights

  • 6-acetylmorphine is the main target metabolite to identify heroin abuse.

  • Morphine, codeine, papaverine and thebaine and metabolites have also been monitored.

  • Metabonomics and the metabolome offers clues for the opiates mechanisms of toxicity.

  • Multiple biomarkers of heroin abuse leads robust forensic and clinical interpretations.

  • The application of multiple omics techniques to toxicology it yet to naive.

Abstract

Opium poppy has important medical, socioeconomic, forensic and political implications. More than 80 benzylisoquinoline alkaloids have been described, many of them with relevant therapeutic properties such as morphine, codeine, papaverine and noscapine. Heroin, a semi-synthetic drug produced from morphine is a worldwide serious cause of morbidity and mortality. Heroin dependence is complex phenomenon with environmental and genetic influence, and several biomarkers of exposure have been proposed. This work aims to review the metabolism and metabolomics of opiates with particular interest on their relevance as potential clinical and forensic antemortem and postmortem biomarkers. It is known that the heroin is mainly a prodrug that is rapidly deacetylated in blood to its active metabolite, 6-acetylmorphine, which is then subsequently slowly deacetylated to morphine. Therefore, 6-acetylmorphine has been used as the main target metabolite to prove heroin abuse in clinical, but mostly in forensic routine. Nevertheless, its applicability is limited due to the reduced detection window. Therefore, morphine (and its metabolites morphine-3-glucuronide and morphine-6-glucuronide), codeine, codeine-6-glucuronide, 6-acetylcodeine, noscapine (and its metabolites meconine, desmethylmeconine, and cotarnine), papaverine (and its metabolites 6-desmethylpapaverine, hydroxypapaverine, dihydroxypapaverine, 6-desmethylpapaverine-glucuronide) and thebaine (and acetylthebaol and the non-acetylated analog thebaol) have been additionally recommended to obtain the most reliable results possible. More recently, the identification by metabolomics analysis of several endogenous compounds offered an alternative approach of significant importance to uncover toxic effects. Profound alterations in the neurotransmitters levels and energy and amino acid metabolism have been reported with l-tryptophan, 5-hydroxytryptamine and 5-hydroxyindoleacetate being suggested as potential non-specific biomarkers of long-term heroin addiction. These endogenous metabolic profiles and exogenous components that together comprise the exposome will certainly help to uncover metabolic disturbances and patterns that may be associate to addiction with relevant clinical and forensic implications.

Introduction

The rapid progress in analytical technology, namely of the “omics sciences” (i.e., transcriptomics, proteomics, lipidomics and metabolomics) lead to the comprehensive analysis of endogenous molecules.1, 2, 3 Metabolomics or metabonomics or metabolic profiling (i.e., comprehensive analysis of the metabolome) is one of the most widely used “omics techniques” and its application has been expanded to various scientific fields namely to toxicology in attempts to elucidate the mechanism of toxicity of xenobiotics by offering “clues” to explain phenotype variation.4 As it is well known, metabolites are produced during different biosynthetic and catabolic pathways that contribute to the complexity of the metabolome. Therefore, metabolomics aims to extract, separate and analyze the totality of small molecules present in any biological sample (i.e., fluids or solid tissues).5 The word “metabolome” (i.e., the total set of endogenous metabolites in an organism) was first coined in 1998 and refers to low-molecular weight molecules (<1500Da) that can be used as biomarkers, such as amino acids, organic acids, fatty acids, sugars and sugar phosphates, and it is positioned at the bottom of “omics cascade” which is the closest link to phenotype.6 Due to its potential to map early biochemical changes in disease, metabolomics provides an opportunity to develop predictive biomarkers of the diagnosis of certain pathological conditions, physiological states and interactions with environmental aspects (i.e., diet, lifestyle, gut microbial activity, and genetics), drug discovery, pharmacometabolomics and personalized medicine.7, 8, 9

The use of the term “biomarker” dates back to 1980.10 In 1998, the National Institutes of Health Biomarkers Definitions Working Group described a biomarker as “a characteristic that is objectively measured and evaluated as an indicator of normal biological courses, pathogenic progressions, or pharmacologic responses to a therapeutic intervention”.10,11 The World Health Organization, defines biomarker as ‘‘any substance, structure or process that can be measured in the body or its products and influence or predict the incidence or outcome of the disease”. Typically, biomarkers can be classified into four types: i) diagnostic biomarkers for a specific disease; ii) prognostic biomarkers to evaluate the clinical outcome; iii) predictive biomarkers to evaluate the response to a particular treatment; and iv) predisposition biomarkers to uncover the inherent or acquired ability of the organism of developing a disease.12,13

Opiates are defined as the derivatives of opium alkaloids, the most relevant representatives being morphine, codeine and heroin. Heroin (i.e., 3,6-diacetylmorphine, diamorphine) is semi-synthesized drug obtained by diacetylation of morphine and has extremely higher potency than morphine due to its higher lipophilicity, which can strengthen its permeability through the blood-brain barrier.14 The drug was firstly synthetized by the chemist Charles Romley Alder Wright in 1874 while boiling morphine with acetic anhydride and then introduced into medicine by Bayer® laboratories in 1898 as a cough suppressant, possessing higher opioid activity comparatively to morphine.15 It was commercialized as Heroin® and the name likely comes from the German word “heroisch” (i.e., something extremely powerful). When its addictive potential was recognized, Bayer® ceased its production in 1913. Nowadays, heroin is the second most popular recreational drug (after Cannabis) and is arguably the world's most widely abused opioid and the most physiologically and psychologically problematic illicit psychoactive substance; indeed more abusers die each year from heroin abuse, and more are forced to seek treatment for addiction, than for any other illicit drug.16 Heroin has also the highest dependence, tolerance and withdrawal score.17 Moreover, there is a tenuous difference between recreational and fatal doses, and variations in street drug purity may cause overdoses. In addition, heroin is the drug most frequently associated with intravenous administrations and therefore there is an increased risk of bloodborne diseases transmission such as human immunodeficiency virus infection and acquired immune deficiency syndrome (HIV/AIDS) and hepatitis B and C.16,18 Heroin has a very short half-life (i.e., approximately 2–4 min) since it is rapidly metabolized to 6-acetylmorphine (6-AM), morphine and other metabolites, meaning that heroin itself cannot be a useful target analyte to identify heroin abuse in practice. Although 6-AM has been claimed to be the most relevant biomarker of heroin abuse, it also has some limitations.19, 20, 21, 22, 23, 24 Therefore, alternative approaches such as metabolomics, by providing information of endogenous metabolic profiling in a global view, may help to reveal heroin abuse and better understand related mechanisms. In this review, metabolomics application to biological samples such as urine, blood or other tissues, focusing on the studies of chronic and acute administration of opiates will be presented together with metabolism and an outline of basic analytical techniques used in metabolomics. Limitations of metabolism and metabolomics studies and future perspectives will be also discussed.

Section snippets

Methodology

An exhaustive computer assisted search of the literature was carried out in PubMed (US National Library of Medicine, Bethesda, MD), Web of Science (Thomson Reuters, Philadelphia, PA), SciFinder (Chemical Abstracts Service, Columbus, OH), Scopus (Elsevier B.V., Amsterdam, the Netherlands), Google Scholar (Google Inc, Mountain View, CA) and books without a limiting period, concerning the metabolomics and metabolism of opiates, namely morphine, heroin (or diamorphine), codeine, papaverine and

Opioids versus opiates

Opioids are substances that interact with opioid receptors.25 Terms such as narcotics, hypnoanalgesics and narcoanalgesics are now considered inadequate since they includes several other xenobiotics that induce sleep. They have also been called opiates, firstly a generic name, but now restricted to naturally occurring alkaloids derived from the annual herbaceous opium poppy plant (Papaver somniferum and Papaver album), namely morphine (10%), codeine (or methylmorphine, 0.5%), papaverine (1%),

Metabolism of heroin, codeine and morphine

Fig. 1 presents the main metabolic routes of heroin, codeine and morphine. Heroin is rapidly either enzymatically or spontaneously metabolized by a sequential hydrolysis/deacetylation in vivo spontaneously to 6-AM and then to morphine.39, 40, 41, 42 Heroin has a very low affinity for μ-opioid receptors and functions mainly as a highly lipophilic prodrug of its active metabolites 6-AM, morphine and morphine-6-glucuronide.14,26,43, 44, 45 The structural modifications of heroin (relative to

Interpretation pitfalls of the opiate toxicological analyses results

There are several factors that may complicate the interpretation of the results of the opiate toxicological analyses, namely: i) codeine contamination of heroin; ii) the presence of morphine and codeine in poppy seeds or medicines for the treatment of pain and cough suppression; iii) the metabolic conversion of codeine and heroin to morphine; iv) the genetic polymorphisms that can affect the metabolism; v) the different rates of urinary excretion of codeine and heroin metabolites; and vi)

Metabolomics of morphine

Morphine treatment lead to profound alterations in the neurotransmitters levels and energy and amino acid metabolism.118,119

Metabolomics of heroin

Previous metabolomics studies in rat's serum and urine samples exposed to heroin have shown metabolic alterations that returned to normal levels after deprivation of the drug.168 Moreover, the reduced weight registered in heroin exposed animals is most probably related to the attempt to maintain energy/ATP levels by168: i) upregulation of tricarboxylic acid cycle as evidenced by the increase of citrate, a crucial intermediate, after heroin administration even for 4 days after withdrawal; and

Analytical techniques in metabolomics

Although various analytical techniques can be used for metabolomics, NMR spectroscopy and mass spectrometry (MS) are the most common categories.168,190,191 NMR-based techniques have been used since the beginning of metabolomics history and have some advantages: i) requires minimal or no need for sample pretreatment before analysis; ii) it is useful for qualitative and quantitative analysis; iii) it is generally a nondestructive technique, allowing intact metabolites to be analyzed even in crude

Chemometric tools in metabolomics

In targeted metabolomics, by using internal standards, especially those isotopically labeled, semi-quantitative and quantitative analysis can be performed.196 This approach can reduce the likelihood of analytical artifacts and the interference by high abundance species by applying specific metabolite extraction techniques.196 However, since untargeted metabolomics provides information of endogenous metabolic profiling in a global view, the number of the detected metabolites ranges from at least

Concluding remarks

Metabolomics is a relatively new but also a rapidly growing technique that provides information of endogenous metabolic profiling in a global view. It may revolutionize knowledge about pathophysiology and diagnosis of diseases, namely uncovering the multiple metabolic changes that may be related to the toxic effects of psychoactive substances. Metabolomics is also a valuable early diagnostic technique to identify patients at increased risk of neuropsychiatric disorders prior clinical symptoms

Disclosure statement

The authors have no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending or royalties. No writing assistance was utilized in the production of this manuscript.

Acknowledgements

This work was supported by grants from CESPU (TramTap-CESPU-2016 and ChronicTramTap_CESPU_2017); and project NORTE-01-0145-FEDER-000024, supported by Norte Portugal Regional Operational Programme (NORTE 2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund. Ricardo Dinis-Oliveira acknowledges Fundação para a Ciência e a Tecnologia for his Investigator Grant (IF/01147/2013).

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