- Background of the study
The study of herbal medicines and the use of plant leaves, stems, roots, seeds and the latex, for human benefits, is an age long event for human benefits (Okafor et al., 1994). Herbal medicine is fast emerging as an alternative treatment to available synthetic drugs for the treatment of diseases possibly due to lower cost, availability, fewer adverse effects and perceived effectiveness (Ubaka et al., 2010). The exploitation of cheap agricultural materials to manufacture industrial products will enhance the development of rural agro-based economy (Kronbergs, 2000; Sain and Panthapulakkal, 2006). The historic role of medicinal plants in the treatment and prevention of diseases and their role as catalysts in the development of pharmacology do not however, assure their safety for uncontrolled use by an uninformed public (Matthews et al., 1999). The use of plants in the management and treatment of diseases started with life. In more recent years, with considerable research, it has been found that many plants do indeed have medicinal values. Some medicinal plants used in Nigeria include Garcina kola, used in the treatment of asthma, Carica papaya, used as a remedy for hypertension, Ocimum basilicum, a cure for typhoid fever, and Cola nitida, for treatment of pile (FAO, 1996). In Nigeria, fermented Prosopis africana seeds are popularly used as food seasoning. It is evident that fermented food condiments are good sources of nutrients and could be used to produce complementary food supplements (Achi, 2005). The food flavouring condiments are prepared by traditional methods of uncontrolled solid substrate fermentation resulting in extensive hydrolysis of the protein and carbohydrate components (Fetuga et al., 1973; Eka, 1980). Apart from increasing the shelf life, and a reduction in the anti-nutritional factors (Odunfa, 1985; Barimalaa et al., 1989; Achi and Okereka, 1999), fermentation markedly improves the digestibility, nutritive value, and flavours of the raw seeds. Fermented products remain of interest since they do not require refrigeration during distribution and storage. The traditional condiments have not attained commercial status due to the very short shelf life, objectionable packaging materials, stickiness and the characteristic putrid odour (Arogba et al., 1995). Fermented condiments often have a stigma attached to them; they are often considered as food for the poor.
Liver damage due to ingestion or inhalation of hepatotoxins such as drugs is increasing worldwide, and conventional drugs used in the management of drug induced liver damage are mostly inadequate and have serious adverse effects (Ozougwu, 2011). In spite of the tremendous strides in modern medicine, there are grossly few drugs that stimulate liver function, offer protection to the liver from damage or help regeneration of hepatic cells. Chronic hepatic diseases is one of the foremost health problems worldwide, with liver cirrhosis and drug induced liver injury accounting for the ninth leading cause of death amongst the western and developing countries population (Mohamed Saleem et al., 2010). About 20,000 deaths are reported every year due to liver disorders (Gupta and Misra, 2006). As said earlier conventional drugs used in the management of drug induced liver damage are mostly inadequate and have serious adverse effects (Ozougwu, 2011). It is, therefore, necessary to explore the herbal options in the management of drug induced liver damage to replace currently used drugs of low efficacy and safety.
1.1 Prosopis africana
1.1.1 Ecological and some pharmacological importance of Prosopis africana seed
Prosopis africana is a leguminous plant of the Fabaceae family. It is a flowering plant that is locally called “kiriya” in Hausa, “okpehe” in Ibo and Idoma and “gbaaye” in Tiv languages of Nigeria. The leaves, branches, bark and roots are used for several purposes in traditional medicines (Kalinganire et al., 2007). Prosopis africana plant is a tropical leguminous tree that is readily distinguished by its dark, pale drooping foliage with small pointed leaflets. The tree is about 12m to 18m high and up to 2.2m in girth. The dry pods which are between 10cm and 15cm long and about 2cm thick contain numerous ellipsoid seeds of about 15 to 18 (Ogunshe et al., 2007).The only known usage of these seeds, presently in Nigeria, is as food seasoning, which is particularly common among the Idomas of Benue State. The seeds are processed in the same way as locust bean seeds. The seeds have protein content of between 39 and 40 per cent (Balogun, 1982). It is traditionally used for formulation of animal feeds and preparation of local condiments through boiling and fermentation processes (Aremu et al., 2006).
The tree is of great economic value to man and animal, it fixes nitrogen to enrich the soil, generates hardy timbers, produces protein rich leaves and sugary pods used as feed stuffs for ruminants (Annongu et al., 2004). However, the disadvantage of Prosopis is the high content of anti-nutritive factor such as tannins, haemagglutinins, prosopine and toxic amino acids which are capable of inducing adverse effect on simple stomached animals when consumed without adequate processing (Cheeke and Shull, 1985). The seeds could be used as a protein supplement for low-protein foods and seeds such as cereal grains for animals (Maragoni and Alli, 1987). The seeds could also serve as a good source of carbohydrate concentrate for all classes of livestock.
1.1.2 Pharmacological Properties of Prosospis africana.
The methanol stem bark extract of Prosopis africana is used for anti-inflammatory and pain relief medicine in humans. Likewise, the tannins and dye in the bark is utilized in the leather industry (Ayanwuyi et al., 2010). The leaves and stem are used for treating toothache. The fruits (pods) are used as fodder for ruminant animals (Amusa et al., 2010). In the middle belt states of Nigeria, fermented Prosopis africana seeds are popularly used as food seasoning. It is a source of low cost protein. Gels that could be used for pharmaceutical tablet formulation is obtained from Prosopis africana gum. The endocarp gum of Prosopis africana seed contains high content of galactose and mannose. Galactose is a special type of natural sugar that gives sustained energy for a longer time compared to other sugar. Mannose is important for treatment of urinary tract infections (Achi and Okolo, 2004). Likewise, the seeds have been reported to have 4445 kcal/kg of food energy which is higher than the 2500 to 3000 kcal/kg daily requirement by humans (Barminas et al., 1998).
According to (Kolapo et al., 2009) the stem and root of P. africana indicates a potential candidate plant parts in dentrifice production. The bark of Prosopis africana in this study is used to treat pile (Lawal et al., 2010).
Anticonvulsant properties of Prosopis africana were seen on strychnine (STR) and pentylenetetrazol (PTZ) induced convulsion. Plant extract were effective against PTZ and STR induced convulsions (Ngo-Bum et al., 2009).
Prosopis africana plant extract was pre-screened and evaluated as anti-trypanosomal agents. The result suggests that the plant with promising bioactivity may possess component that may provide the chemical lead towards the discovery of new generation trypanocides that are more potent and less toxic than the currently available and marketed trypanocidal drugs (Osho and Lajide, 2012).
Prosopis africana belonging to the family Fabaceae is being used traditionally as medicine in many African homes. This includes the leaves used in treatment of headache and toothache; leaves and bark are combined in the treatment of rheumatism, skin disease and eyewashes; the roots are used as diuretic, and in the treatment of dysentery, bronchitis and stomach cramps (Gilbert and Neil, 1986; Arbonnier, 2002). The prosopis gum has been used in the present day research as bio-adhesive agent in delivery of metformin, this show a synergistic effect (Adikwu and Nnamani, 2005).
Prosopis gum can be used to treat infection, skin irritation and in the management of wound. These studies suggest that a mixture of bovine mucin, cicatrin and prosopis gum has a better healing effect than cicatrin powder alone (Momoh et al., 2008). According to Ojo et al. (2006), Prosopis africana leaf extract was observed to have hepatoprotective potentials, this could be related to the high concentration of tannin in the leaf.
1.1.3 Scientific Classification of Prosopis africana Seed
Source: (Achi and Okereka, 1999)
Fig. 1: Prosopis africana seed
Fig. 2: Fermented seed of Prosopis africana
Liver is a self regenerating organ that plays important roles in the body. It functions not only in metabolism and removal of exogenous toxins and therapeutic agents responsible for metabolic derangement but also in the biochemical regulation of fats, carbohydrates, amino acids, protein, blood coagulation and immunomodulation function (Ram and Goel, 1999). Due to its ability to regenerate, even a moderate cell injury is not reflected by measurable change in its metabolic function. However, damage caused by lipid peroxidation on the membrane of the hepatocytes allows the leakage of some cytoslic enzymes of the liver into the blood stream (Plaa and Hewitt, 1982).
1.2.1 Epidemiology and Statistics of Drug Induced Liver Injury.
Drug induced liver damage is a health problem worldwide and is expected to increase as the number of drugs being consumed increases. It is a major health issue that challenges not only health care professionals but also the pharmaceutical industry and drug regulatory agencies (Saleem et al., 2008). Drug induced liver injury is the most commonly cited reason for withdrawal of already approved drugs from the market (Butura, 2008). According to the United States Acute Liver Failure Study Group, drug-induced liver injury accounts for more than 50% of acute liver failure, with hepatotoxicity caused by overdose of paracetamol accounting for 39% and idiosyncratic liver injury triggered by other drugs accounting for about 13% (Holt and Ju, 2006). Drug-induced liver toxicity accounts for approximately half of the cases of acute liver failure and mimics all forms of acute and chronic liver disease (Kaplowitz, 2001). The reported incidence of anti-tuberculosis drugs induced hepatotoxicity indicated that the developing countries are having difficulties in systematic steps for prevention and management of tuberculosis drugs induced hepatotoxicity. Despite the frequency of drug induced liver injury being relatively low, data from the centers for disease control and prevention in the U.S reported approximately 1600 new acute cases of liver failure annually, of which paracetamol hepatotoxicity accounts for approximately 41% (Norris, and Lewis, 2008). The most commonly implicated drugs involved in acute liver injury are summarised in Table 1.
Table 1: Commonly reported drugs associated with drug induced liver injuries.
|Non-Steroidal Anti-Inflammatory Drugs|
(Isoniazid, Rifampicin, Pyrazinamide)
Anti-Retroviral Drugs (E.g Ritonavir)
Source: (Chau, 2008)
1.3 Acetaminophen (Paracetamol)
Acetaminophen is an effective antipyretic and analgesic, but its anti-inflammatory properties are minimal, especially compared with non-steroidal anti-inflammatory drugs (NSAIDs). Nevertheless, acetaminophen is preferred over NSAIDs in some patients because it carries a lower risk of gastrointestinal toxicity (eg, ulceration, bleeding) and so may be better tolerated (Burke et al., 2006). It is one of the most important drugs used for the treatment of mild to moderate pain when an anti-inflammatory effect is not necessary (Nwachukwu, 2006). Acetaminophen structure is shown below in Fig. 3.
HO N C CH3
Fig. 3: N-acetyl-p-aminophenol (Acetaminophen)
1.3.1 History of paracetamol
Its history says that when Cinchona tree became scarce in the 1880s, people began to look for alternatives. Two alternative antipyretic agents were developed in 1880s; Acetanilide in 1886 and Phenacetin in 1887. Harmon Northrop Morse first synthesized paracetamol via the reduction of p-nitrophenol with Tin in glacial acetic acid in 1878; however, paracetamol was not used in medical treatment for another 15 years. In1893, Paracetamol was discovered in the urine of individuals that had taken Phenacetin and was concentrated into white crystalline compound with a bitter taste. In 1899, paracetamol was found to be a metabolite of acetanilide. This discovery was largely ignored at that time. In 1948, Brodie and Axelrod determined that the analgesic effect of acetanilide was due to its active metabolite paracetamol. The product was then first sold in 1955 by McNeil laboratories as a pain and fever reliever for children, under the brand name Tylenol children’s elixir (Vidhya and Metillda, 2012).
1.3.2 Pharmacokinetics of Paracetamol
Paracetamol is administered orally. Absorption is related to the rate of gastric emptying and peak blood concentrations are usually reached in 30-60 minutes. Acetaminophen is slightly bound to plasma proteins and is primarily metabolized by hepatic microsomal enzymes and converted to acetaminophen sulphate and glucoronide, which are pharmacologically inactive. Less than 5% is excreted unchanged. A minor but highly active metabolite (N-acetyl-p-benzoquinone) is important in large doses because of its toxicity to both liver and kidney (Prescott et al., 2006). The half life of acetaminophen is 2-3 hours and is relatively unaffected by renal function.
1.3.3 Intravenous and Oral Administration of Paracetamol
Paracetamol has previously been available for intravenous use in the form of its pro-drug, propacetamol. Used in France since 1985, propacetamol, provided as a powder for reconstitution, is water soluble and rapidly hydrolysed by plasma esterases to form paracetamol and diethylglycine; a dose of 1 g propacetamol provides 0.5 g paracetamol after hydrolysis. In a study of patients undergoing dental extraction, propacetamol was significantly better than placebo for all measured parameters; pain relief, pain intensity, patient’s global evaluation and duration of analgesia (Moller et al., 2005). Advantages of intravenous paracetamol over not associated with pain on injection or contact dermatitis. Paracetamol is bioequivalent to propacetamol (Flouvat et al., 2004).
In a study of 35 patients undergoing day-surgery, intravenous propacetamol (the IV prodrug of Paracetamol) reached therapeutic plasma concentrations more quickly and predictably than oral Paracetamol (Holmer-Pettersson et al., 2004). Paracetamol plasma concentrations were observed for the first 80 minutes after administration of either 1 g or 2 g oral Paracetamol or 2 g intravenous propacetamol. Intravenous paracetamol provided an average concentration within the therapeutic range after 20 minutes. There was a large and unpredictable variability with oral administration; some patients who received 1 g orally did not achieve detectable plasma levels within the 80 minute study period, and the average plasma concentration after receiving this dose was sub therapeutic throughout. 2 g oral paracetamol achieved a median plasma concentration within the therapeutic range after 40 minutes, suggesting that when paracetamol is given orally, a loading dose can reduce the time needed to achieve therapeutic levels.
1.3.4 Metabolism of Paracetamol
Paracetamol is metabolised in the liver via three pathways – glucuronidation, sulphation (both account for 95% of metabolism) and cytochrome P450 system (5%). Acetaminophen is metabolically activated by cytochrome P450 to form a reactive metabolite that covalently binds to protein (Mitchell et al., 1973). The reactive metabolite was found to be N-acetyl-p-benzoquinone imine (NAPQI), which is formed by a direct two-electron oxidation (Dahlin et al., 1984). NAPQI is detoxified by glutathione (GSH) to form an acetaminophen- GSH conjugate. After a toxic dose of acetaminophen, total hepatic GSH is depleted by as much as 90%, and as a result, the metabolite covalently binds to cysteine groups on protein, forming acetaminophen- protein adducts (Mitchell et al., 1973). This mechanism is shown in Fig. 4.
Fig 4: Schematic representation depicting the role of metabolism of acetaminophen toxicity
Source: (James et al., 2003)
Paracetamol is rapidly absorbed from the small intestine. Peak serum concentrations occur within 1-2 hours for standard tablet or capsule formulations and within 30 minutes for liquid preparations. Peak serum concentrations after therapeutic doses do not usually exceed 130nmol/l (20gm/l) (Leshna et al., 1976).
Twenty percent (20%) of the ingested dose undergoes first-pass metabolism in the gut wall (sulphation). Distribution is usually within 4 hours of ingestion for standard preparations and 2 hours for liquid preparation. Volume of distribution is 0.91/kg2. Further elimination occurs by hepatic biotransformation. After therapeutic doses, the elimination half-life is 1.5-3 hours (Nahid et al., 2005). Over 90.6% is metabolised to inactive sulphate and glucuronide conjugates that are excreted in the urine. Metabolism of the remainder is via cytochrome P450 and results in the highly reactive intermediary compound N-acetyl-p-benzoquinone imine (NAPQI). In normal conditions, NAPQI is immediately bound by intracellular glutathione and eliminated in the urine as mercapturic adducts (Daly et al., 2008). With increased paracetamol doses, greater production of NAPQI may deplete glutathione stores. When glutathione depletion reaches a critical level (about 30% of normal stores), NAPQI binds to other proteins, causing damage to the hepatocyte. Glutathione depletion itself may be injurious (Kupeli et al., 2006).
The commonest target organ in paracetamol poisoning is the liver and the primary lesion is acute centrilobular hepatic threshold. In adults, the single acute threshold dose for severe liver damage is 150-250mg/kg though there is marked individuals variation in susceptibility (Prescott et al., 2006).
1.3.5 Pharmacodynamics of Paracetamol
The major active metabolites of paracetamol are sulphates and glucuronide conjugates. Its main mode of action is to inhibit the activity of the enzyme cyclooxygenase (COX) (James et al., 2003). COX enzymes are necessary for the production of prostaglandins. Prostaglandins are a form of hormone (although rarely classified as such) that are indicated to be mediators of pain, fever and inflammation. The half-life of paracetamol may be measured either by salivary or by plasma counts. Both measurements give varying half-life between 1 and 4 hours (Lee et al., 1996). Peak levels are reached 40-60 minutes after ingestion. It has been proposed that paracetamol aids in the reduction of pain by increasing serotonergic neurotransmissions (Garrone et al., 2007).
1.3.6 Paracetamol Hepatotoxicity
Overdose of paracetamol leads to ‘paracetamol hepatotoxicity,’ which mainly results into liver injury but is also one of the most common causes of poisoning all over the world (Norris, and Lewis, 2008). Many people who develop paracetamol toxicity may feel no symptoms at all in the first 24 hours that follow overdose of paracetamol. Others may initially experience nonspecific complaints like vague abdominal pain and nausea. As the paracetamol toxicity increases, signs of liver failure like low blood sugar; low blood pH, easy bleeding, and hepatic encephalopathy may develop. Timely treatment can cure the condition of the patient but untreated cases may result in death. Often a liver transplant is needed if damage to the liver gets severe. The risk of paracetamol toxicity increases with excessive alcohol intake, fasting or anorexia nervosa, and also with the use of certain drugs like isoniazid (Vidhya and Metillda, 2012).
Events that produce hepatocellular death following the formation of acetaminophen protein adducts are poorly understood. One possible mechanism of cell death is that covalent binding to critical cellular proteins results in subsequent loss of activity or function and eventual cell death and lysis. Primary cellular targets have been postulated to be mitochondrial proteins, with resulting loss of energy production, as well as proteins involved in cellular ion control (Nelson, 1990). Tirmenstein and Nelson (1989) and Tsokos-Kuhn et al. (1988) reported alterations of plasma membrane ATPase activity following toxic doses of acetaminophen.
In addition to hepatotoxicity, NAPQI inhibits mitochondrial respiration by blocking electron transport between the cytochrome b/c complex and the cytochrome oxidase complex within the electron transport chain (Porter and Dawson, 1979). Fasting is a risk factor, possibly because of depletion of hepatic glutathione reserves (Nolan et al., 1994). Chronic alcoholism which also induces CYP2E1 is also well known to increase the risk of paracetamol induced hepatotoxicity (Nwodo et al., 2010).
In normal doses, paracetamol does not irritate the lining of the stomach or affect blood coagulation, the kidney or the fetal ductus arteriosus (as NSAIDs can) like NSAIDs and unlike opiod analgesics paracetamol has not been found to cause euphoria or alter mood in any way. Since this molecule is achiral, it does not have a specific rotation (Prescott et al., 2006). The words acetaminophen and paracetamol are both derived from the chemical names for the compounds N acetyl-para-aminophenol and para-acetyl animo-phenol respectively.
184.108.40.206 Paracetamol Hepatotoxicity and Alcohol Consumption
It is claimed that chronic alcoholics are at increased risk of paracetamol (acetaminophen) hepatotoxicity not only following over dosage but also with its therapeutic use. Increased susceptibility is supposed to be due to induction of liver microsomal enzymes by ethanol with increased formation of the toxic metabolite of paracetamol. However, the clinical evidence in support of these claims is anecdotal and the same liver damage after overdosage occurs in patients who are not chronic alcoholics. Many alcoholic patients reported to have liver damage after taking paracetamol with ‘therapeutic intent’ had clearly taken substantial overdoses (Vidhya and Mettilda, 2012).
The paracetamol-alcohol interaction is complex; acute and chronic ethanol has opposite effects (Garry et al., 2004). In animals, chronic ethanol causes induction of hepatic microsomal enzymes and increases paracetamol hepatotoxicity as expected (ethanol primarily induces CYP2E1 and this isoform is important in the oxidative metabolism of paracetamol). However, in man, chronic alcohol ingestion causes only modest (about two fold) and short-lived induction of CYP2E1, and there is no corresponding increase (as claimed) in the toxic metabolic activation of paracetamol. Acute ethanol inhibits the microsomal oxidation of paracetamol both in animals and man. This protects against liver damage in animals and there is evidence that it also does so in man. The protective effect disappears when ethanol is eliminated and the relative timing of ethanol and paracetamol intake is critical (Vidhya and Mettilda, 2012).
Hepatotoxicity from therapeutic doses of paracetamol is unlikely in patients who consume moderate to large amounts of alcohol daily. However, patients with severe alcoholism should be instructed or supervised about the correct dosage of paracetamol. The depression often associated with alcoholism may make them more likely to take an overdose of paracetamol (Garry et al., 2004).
In many of the reports where it is alleged that paracetamol hepatotoxicity was enhanced in chronic alcoholics, the reverse should have been the case because alcohol was actually taken at the same time as the paracetamol. Chronic alcoholics are likely to be most vulnerable to the toxic effects of paracetamol during the first few days of withdrawal but maximum therapeutic doses given at this time have no adverse effect on liver function tests. Although the possibility remains that chronic consumption of alcohol does increase the risk of paracetamol hepatotoxicity in man (perhaps by impairing glutathione synthesis), there is insufficient evidence to support the alleged major toxic interaction (Prescott, 2000). Chronic consumption of alcohol for three consecutive days may cause inflammation and scarring of the liver cells depending on diet, sex, immune status, gut flora and the capacity of the metabolising enzymes (Nwodo, 2012).