ATTENTION:

BEFORE YOU READ THE ABSTRACT OR CHAPTER ONE OF THE PROJECT TOPICS BELOW, PLEASE READ THE INFORMATION BELOW.THANK YOU!

INFORMATION:

YOU CAN GET THE COMPLETE PROJECT OF THE TOPIC BELOW. THE FULL PROJECT COST N5,000 ONLY. THE FULL INFORMATION ON HOW TO PAY AND GET THE COMPLETE PROJECT IS AT THE BOTTOM OF THIS PAGE. OR

YOU CAN CALL: 08068231953, 08137701720

WHATSAPP US ON: 08137701720

THE EFFECT OF CRUDE OIL CONTAMINATED WATER ON CONTRACTILE ACTIVITY OF THE TRACHEA SMOOTH MUSCLE IN ADULT MALE WISTAR RAT

ABSTRACT

Increased blood pressure has been associated with exposure to petroleum products by inhalation or administration of contaminated water. The mechanism of action has however not been elucidated. This study was undertaken to investigate the effect of petroleum products on cardiovascular functions using three different routes of administration and the possible involvement of oxidative stress in the mechanism of action. Sprague Dawley rats were divided into four main groups; control, diesel (automotive gas oil), kerosene (dual purpose kerosene) and petrol (premium motor spirit). Each of the groups except control was subdivided into three; ingestion, inhalation and water contaminated groups; with 10 rats in each group and sub-group. The control was not exposed to any treatment. The diesel, kerosene and petrol sub-groups, were exposed to their corresponding contaminants via ingestion, inhalation and water contamination respectively. Each administration and exposure lasted eight weeks. The results showed that blood pressure and heart rate were significantly increased (p<0.05) in the treated groups when compared with control. However, there was insignificant reduced blood pressure in petrol ingestion group. The significant increase in the blood pressure and heart rate persisted one week after stoppage of exposure in all the groups; suggesting that exposure to petroleum products could cause hypertension; the effect could be as a result of the ability of petroleum products to cause sensitization of the vascular smooth muscle to catecholamines, elicited by impaired endothelium-dependent and -independent vasomotor function. Baroreflex response was significantly (p<0.05) increased in diesel and kerosene groups compared with control, however the increase in the petrol was only significant for a while and returned to the control level by 25 seconds. This suggests that with exposure to petrol the baroreceptors still reset the blood pressure to a new higher than normal value as though normal. The activities of baroreceptors have been altered by diesel and kerosene such that they could no longer reset the arterial blood pressure. Exposure to diesel was most deleterious in all except in the inhalation group. There was reduced body weight gain (p<0.05) in all exposed rats, which was most severe in the diesel group, when compared with control as well as kerosene and petrol groups. Exposure to petroleum products dissolved fat and lipid in the body causing degeneration of fat store. The reduced weight gain might also be associated with decreased food intake seen in the study especially the diesel group. Anaemia was implicated as there were significant (p<0.05) reductions in RBC, PCV, and Hb. Platelets and lymphocyte reduced significantly in diesel and petrol groups but not significant in kerosene groups. A significant increase in WBC was recorded in diesel, kerosene and petrol groups. Liver damage as a result of increased reactive oxygen species was more severe in the diesel ingestion group as Alanine amino transferase (ALT), Aspartate amino transferase (AST) and Alkaline phosphatase (ALP) increased significantly p<0.05) in all the groups. The results also showed significant increase in lipid peroxidation, as concentration of MDA increased significantly (p<0.01) in all the treated groups. Catalase

(CAT), Glutathione reductase (GSH) and superoxide dismutase (SOD) activities were (p<0.05) reduced significantly in serum and tissue homogenate at varying proportions in the different groups. In the urine samples there was significant reduction in creatinine and urea value compared with control, while creatinine and urea in serum increased significantly suggesting renal function impairment. In conclusion, exposure to petroleum products resulted in increased blood pressure and heart rate; the baroreceptors were impaired by diesel and kerosene; caused anaemia, reduced platelets and lymphocytes and caused renal dysfunction. The severity of the effects was most severe in the diesel groups. This study suggests that afore-mentioned observations are partly caused by oxidative stress by altering the levels of CAT, GSH, SOD and MDA.

CHAPTER ONE

1.0 INTRODUCTION

1.1 BACKGROUND TO THE STUDY

The primary cardiac function is to impart energy to blood in order to generate and sustain an arterial blood pressure necessary to provide adequate perfusion of organs. Cardiovascular dysfunctions such as hypertension, stroke, and coronary heart disease are associated with many underlining disease conditions and environmental factors. Hypertension is the most prevalent of all cardiovascular events (Kaplan, 2006; Julia et al., 2012). Petroleum products are frequently used in our environment and are ubiquitous environmental toxicant; which affect many physiological parameters and cause stress in man and animals (Ovuru and Ekweozor, 2004). Interest on the adverse effects of petroleum hydrocarbons has grown in recent years, and focus has been on the deleterious effects of these products on various systems in the body. Exposure could be by ingestion of food crop from polluted soil and contaminated sea foods, inhalation of vapour in the environment, (from vehicle exhaust, as petrol attendant or refinery worker) or drinking contaminated water. Contamination with petroleum products is increasing both in rural and urban areas. Acute exposure also occurs following spillage, blowouts, pipeline-vandalization and tanker accidents. There is increased use of petroleum products for power generation in domestic and industrial environment (Anigbogu and Ojo, 2009). Diesel, gasoline and kerosene are among the commonly used fractionated products of crude petroleum. These fractions contain aliphatic, aromatic and a variety of other branched saturated and unsaturated hydrocarbons at variable concentrations. The constituents of the vapours from these fractions, to a greater extent, depend on the composition of their liquid forms and carbon content, which varies with the brand and storage period (EHC 20, 1982; Henderson et al., 1993; Kato et al. 1996; Anderson et al., 1995).

Petroleum products are integral part of our modern lives. It is almost impossible to avoid exposure to hydrocarbon from petroleum products, whether it is from gasoline fumes at pumps, spilled crankcase oil on asphalt solvent used at home or work or pesticide applications that use petroleum product as carrier. Petroleum products are used in the domestic environment and are still frequently stored in the homes or garage in unmarked container or beverage bottles. They may also have attractive aromas and be brightly coloured. It is not surprising therefore that most cases of exposure in the homes involve accidental ingestion by young children/adults. A larger proportion of childhood poisoning occurs in the pre-school group. Children under 6 years accounted for more than 50% of all exposures reported to poisons centre in the USA in 1996, of which 2.4% involved hydrocarbons (U.S. EPA 2009). Contamination with petroleum product is not only limited to the marine environment. Pollution at drill sites and oil spills on land used for agricultural purposes, as well as petroleum or diesel waste; pose serious risks exposure to occupational public, terrestrial wildlife, mammals and livestock raised on these lands (Igwebuike et al., 2007). Some components of petroleum have the potential to bioaccumulate within susceptible aquatic organisms and thus passed by trophic transfer to other levels in the food chain (Gardner et al., 1991).

The adverse effects of bioaccumulation in exposure to petroleum products are manifold and vary depending on the concentration of the substances and the length of time that one is exposed. Breathing petroleum vapour can cause nervous system defects such as

headache, nausea, dizziness and respiratory irritation (Steffe et al., 1996).

Very high exposure to petroleum products can cause coma and death (Levy and Pappano, 2007). Liquid petroleum products which come in contact with the skin can cause irritation and some are absorbed through the skin. Chronic exposure to petroleum products may affect the nervous system, blood and kidneys (Levy and Pappano, 2007). Gasoline is known to contain small amounts of benzene, a known human carcinogen

(Periago and Prado, 2005).

The toxicity of petroleum product is related to its hydrophobicity (Freedman, 1995) because lipid solubility is an important factor in the passage of petroleum components through the plasma membrane of cell as well as the degree of membrane damage (Igwebuike et al., 2007). The most common sources of petroleum contamination can either be from stationary petroleum storage systems or when released to the environment. Contamination can occur from a storage system through;

Leaks in pipes and joints
Leaks from corroded tanks
Over fills and spills while filling tanks

When petroleum products are released into the environment, the petroleum and the products can contaminate the soil, groundwater, surface water and air. Explosive vapours from discharged petroleum products can accumulate in confined space such as an abandoned tank, a subsurface cable vault, and sewer or beneath buildings.

1.2 Major Petroleum Products

1.2.1 Diesel

Diesel, called automotive gas oil (AGO) is produced by fractional distillation of petroleum 220-250^oC (392° F-662° F), typically contains 15-25 carbon atoms. Chemical formula for common diesel fuel is C₁₅H₂₃. There are alternatives that are not derived from petroleum, such as biodiesel , biomass to liquid (BTL) or gas to liquid (GTL) diesel, are increasingly being developed and adopted. To distinguish these types, petroleum-derived diesel is increasingly called petrol diesel. In the UK, diesel fuel for on-road use is commonly abbreviated DERV, standing for DieselEngined Road Vehicle, which carries a tax premium over equivalent fuel for non-road use. Diesel has a higher density than gasoline and is simpler to refine from crude oil. It is most commonly used in transportation in a diesel engine. The generated heat (J/kg) obtained by the burning of diesel in air is 13762J/kg. Petroleum-derived diesel is composed of about 75% saturated hydrocarbons and 25% aromatic hydrocarbons . The hazard experience with petro diesel is that when it spilled on a road, will stay there until washed away by sufficiently heavy rain, whereas gasoline will quickly evaporate. A diesel engine is an internal combustion engine , developed by Rudolf Diesel in 1893 (ATSDR, 1995). Synthetic diesel can be produced from any carbonaceous material, including biomass, biogas, natural gas, coal and many others. The raw material is gasified into synthesised gas, which after purification is converted by the Fischer –Tropsch process to a synthetic diesel. Fatty-acid methyl ester (FAME), perhaps more widely known as biodiesel, is obtained from vegetable oil or animal fats (biolipids ) (Demirbas, 2008); which have been trans –esterified with methanol. It can be produced from many types of oils, the most common being rapeseed oil (rapeseed methyl ester, RME) in Europe and soybean oil (soy methyl ester, SME) in the USA. (Torgov et al., 1994), Diesel displaced coal and fuel oil for steam-powered vehicles in the latter half of the 20th century, and is now used almost exclusively for the combustion engines of self-powered rail vehicles (locomotives and railcars).

1.2.2 Kerosene

Kerosene called dual purpose kerosene (DPO) because it is most commonly used as jet fuel and as heating fuel. It is a thin clear liquid formed from a complex mixture of hydrocarbon (C₁₄H₃₀) with density of 0.78 – 0.81/cm³. Kerosene was first described by al –Razi (Rhazes) as a distillation of petroleum in 9th-century Baghdad . It is obtained from fractional distillation of crude oil between 150 and 270^oC resulting in a mixture of carbon chain that typically contain between 12 and 15 carbon atom per molecule (Ofusori et al., 2009). In the United

Kingdom, kerosene is also known as paraffin. The word kerosene comes from the Greek word ―keros‖ meaning wax. The lethal dose (LD₅₀) of kerosene for 70 kg adult is 100 ml (Patel et al., 2004).

In the early 21st century, kerosene was used to power New York City transit buses. Now, kerosene is used as fuel in portable stoves, kerosene space heaters, and in liquid pesticides. It is the major domestic cooking fuel in many Nigerian homes. It is called dual purpose kerosene because of its dual-use as a fuel that is both “Motor vehicle and non-road, locomotive or marine. A short, one-off exposure to kerosene is unlikely to result in any longterm effects. However, a severe form of lung injury called pneumonitis may occur if liquid kerosene is aspirated directly into the lungs, for example, whilst manually siphoning a tank or from inhaling vomit after swallowing kerosene toxicity is principally due to pulmonary complications if it is inhaled while being ingested (aspiration). Although kerosene is not poisonous, medical advice should be obtained immediately when swallowed as there is a risk of short term lung damage if vomiting occurs. Frequent skin exposure may lead to skin damage (dermatitis); there is a threshold level of exposure to kerosene above which adverse health effects evolves. Breathing large quantities of kerosene vapour or drinking kerosene based liquids may cause non-specific signs such as dizziness, headache, and vomiting

(Oyekale et al., 2012; Mahdi, 1988).

1.2.2.1 Kerosene Toxicological overview

Health Effects of Chronic / Repeated Exposure

The most common health effect associated with chronic / repeated kerosene exposure is Dermatitis (Ritchie et al., 2003) which may be associated with insufficient or inappropriate use of personal protective equipment (PPE) in occupational environments. Lung effects (such as dyspnoea) have been reported, but tend to be associated with ―high level‖ exposures (Ritchie et al., 2003). It is conceivable that similar lung and skin effects may be observed in some individuals following a single, acute exposure

Dermal / ocular exposure

Dermatitis was observed in mice topically exposed to kerosene (applied in muslin cloth) for

15 to 60 minutes each day for one week which resolved within three weeks (Upreti, et al., 1989) Pathological changes (hyperplasia and visual scores of irritation) were also observed in mice exposed twice a week for two weeks to deodorised kerosene, but the lesion severity did not correlate with tumour-promoting activity when compared against four other petroleum products (Walborget al., 1998) When applied three times a week for up to six weeks, repeated cycles of necrosis and regeneration were observed that were deemed sufficient to represent an epigenetic mechanism for tumourigenesis.

Pulmonary pathology (inflammatory cell infiltrates and morphological changes to tracheal epithelia) and cardiovascular changes (resembling atherosclerosis) have been observed in guinea pigs following exposure to high concentrations (up to 34 g m-3) of kerosene aerosol for 15 minutes per day over a three week period(Noa et al., 1987, 1985,1984). Continuous (3 month) exposure of rats and mice to up to 1 g m-3 aviation fuel (JP-8) vapour resulted in male rat-specific pathology (nephropathy) that was not thought to be of relevance to humans (Mattieet al., 1991). In a study carried out by Mann et al., (1977), absorption of kerosene was investigated in primates, the result showed that primates absorbed kerosene from the gastrointestinal tract, but the volumes are very small and do not cause gross neurologic signs. Lee and colleagues (2000), in an experiment found out that cleaning with kerosene resulted in mean carcinogen-DNA adduct levels in the lung which were significantly higher than even the positive controls, regardless of cleaning time. Garciá Mesa and his colleagues in another experiment discovered that the subchronic exposure to vapors of kerosene or its combustion fumes, induced an increase in the activity of lysosomal enzymes in lungs which can be an explanation of the inflammatory response induced in lungs by this agent.

1.2.3 Petrol

Petrol, called premium motor spirit (PMS) gasoline, or petroleum-derived liquid mixture was primarily used as fuel in internal combustion engines. It is obtained from fractional distillation of crude oil between 20^oC – 60^oC, contains 5-11 carbon atom. Easily vapourized, highly flammable and easily ignited. It is also used as a solvent, mainly known for its ability to dilute paint. Exposure to petrol vapour in confined or poorly ventilated areas may cause rapid onset of unconsciousness (Pino et al., 2004). The generated heat (J/kg) obtained by the burning of gasoline in air is 12,200 J/kg. The energy Density depends on the grade and source.

1.2.3.1 Petrol Toxicological Overview

Petrol is a complex mixture of aliphatic and aromatic hydrocarbons derived from blending fractions of crude oil with brand-specific additives. The actual composition of petrol will vary according to the source of crude oil, the manufacturing process and between batches. As with other hydrocarbon solvents, petrol has anaesthetic (narcotic) properties (Table 2). Petrol also contains a number of potentially neurotoxic chemicals including n-hexane, benzene, butadiene, toluene, ethylbenzene, xylene and trimethyl pentane (Ritchie et al., 2001). The approximate concentration of each constituent in liquid petrol and vapour are given in the table below.

Table 1: Average concentration of potentially neurotoxic constituents of liquid petrol and vapour.

	Concentration %w/w)
Chemical	Liquid	Vapour
Benzene	2.5 (0.2 – 4.7)	1.77 (0 – 5.4)
1,3-Butadiene	<0.1	0.65 (0 – 4.6)
Ethylbenzene	2.6 (1.0 – 5.4)	0.009 (0 – 0.1)
n-Hexane	2.5 (0.8 – 5)	1.37 (0 – 6.5)
Toluene	11.4 (2.7 – 21.0)	1.63 (0 – 7.1)
Xylene	10.6 (5.8 – 15.8)	0.48 (0 – 2.1)

Cairney et al., 2002; Cecil et al., 1997)

Numbers in brackets refer to range of values. Vapour values expressed as percentage of total hydrocarbons recovered from air samples obtained during the manual filling of cars (conditions and duration not reported)

General toxicity

Dysfunction of the central nervous systemis the predominant pathological condition associated with chronic exposure to high levels and such effects arising from frequent, recreational exposure (‗sniffing‘ or ‗huffing‘) have been extensively documented (Edminster and Bayer, 1985; Cairney, 2002; Flanagan, and Ives, 1994; Cairney et al., 2004,2005). There is currently insufficient evidence to unequivocally link chronic (occupational) exposure to petrol with other pathological conditions (Kovarik, 2005). This may be because petrochemical workers are potentially exposed to a wide range of chemicals in addition to other confounding factors (IARC,1989). Historically, lead has been identified as the principal component of petrol responsible for neurotoxicity (Kovarik, 2005),and studies have demonstrated a link between lead body burden and neurological deficits as a result of petrol abuse (‗sniffing‘ or ‗huffing‘) (Seshia, 1978). However, it should be noted that the volatility of tetraethyl lead (TEL) is relatively low (0.4 mm Hg at 25ºC) and so prolonged dermal exposure associated with the practice of petrol sniffing is likely to be the predominant route of entry for TEL rather than inhalation (Toxicology update, 1989). Since 2000, petrol has only been commercially available in ‗unleaded‘ form within the UK and most of Europe. This policy limits the concentration of lead in marketable petrol to less than 0.005 g L-1as defined at Annex I of the 1998 EU Directive (EU, 1998). Whilst there is a known association between chronic petrol exposure and renal cancer in male rats (Halder et al., 1985; Olson et al., 1987; Short et al., 1987), there is currently no evidence to link petrol exposure and renal cancer in humans (Trump et al., 1984; IARC, 1989). It is generally accepted that the susceptibility of male rats is mediated via a specific protein (α-2-microglobin) which is absent in other mammals (Olson et al., 1990, 1987).

1.3 Octane Rating

Octane rating or octane number is a standard measure of the performance of a motor or aviation fuel. The higher the octane number, the more compression the fuel can withstand before detonating. In broad terms, fuels with a higher octane rating are used in highcompression engines that generally have higher performance. In contrast, fuels with low octane numbers (but high cetane numbers ) are ideal for diesel engines. Use of gasoline with low octane numbers may lead to the problem of engine knocking (Werner et al., 2007).

1.3.1 Measurement methods

Research Octane Number (RON) is the most common type of octane rating worldwide. RON is determined by running the fuel in a test engine with a variable compression ratio under controlled conditions, and comparing the results with those for mixtures of iso-octane and nheptane. Motor Octane Number (MON) or the aviation lean octane rating, is a better measure of how the fuel behaves when under load, as it is determined at 900 rpm engine speed, instead of the 600 rpm for RON (Werner et al., 2007). MON testing uses a similar test engine to that used in RON testing, but with a preheated fuel mixture, higher engine speed, and variable ignition timing to further stress the fuel’s knock resistance. Depending on the composition of the fuel, the MON of a modern gasoline will be about 8 to 10 points lower than the RON, however there is no direct link between RON and MON. Normally, fuel specifications require both a minimum RON and a minimum MON

Anti-Knock Index (AKI): In most countries, including Australia and all of those in Europe, the “headline” octane rating shown on the pump is the RON, but in Canada, the United States and some other countries, like Brazil, the headline number is the average of the RON and the MON, called the Anti-Knock Index (AKI, and often written on pumps as (R+M)/2). It may also sometimes be called the Pump Octane Number (PON), because of the 8 to 10 point difference noted above, the octane rating shown in Canada and the United States is 4 to 5 points lower than the rating shown elsewhere in the world for the same fuel.

Observed Road Octane Number (RdON), is derived from testing gasolines in real world multi-cylinder engines, normally at wide open throttle. It was developed in the 1920s and is still reliable today. The original testing was done in cars on the road but as technology developed the testing was moved to chassis dynamometers with environmental controls to improve consistency. The selection of octane ratings available at the pump can vary greatly from region to region, for example in Saudi Arabia: Two types of fuel are available at all gas stations in Saudi Arabia. “Premium 91” (RON 91) where the pumps are coloured green, and

“Super Premium 95” (RON 95) where the pumps are coloured red. In United Kingdom: ‘regular’ petrol has an octane rating of 95 RON, with 97 RON fuel being widely available as the Super Unleaded. Tesco and Shell both offer 99 RON fuel. In United States: octane rating is displayed in AKI. In the Rocky Mountain (high elevation) states, 85

AKI (90 RON) is the minimum octane, and 91 AKI (95 RON) is the maximum octane

available in fuel (Werner et al., 2007).

1.4 Inhalant abuse

Inhalants are broad range of drugs whose volatile vapours are taken in via the nose and trachea. They are taken in by volatalization. While some drugs are used for medical purposes, as in the case of nitrous oxide (a dental anxiolytic), this article focuses on inhalant abuse as recreational drugs that are used for their intoxicating effect. Inhaling volatile substances because of their intoxicating effect is called huffing. Most inhalant drugs that are used nonmedically are ingredients of household or industrial chemical products that are not intended to be concentrated or inhaled. Examples include: Hydrocarbons: gasoline, kerosene, butane, Toluene, xylene. Others include: glue, photocopier fluid, aerosol, paint thinner, cleaning and lighter fluids. The recreational use of inhaling hydrocarbons and other volatile solvents for the purposes of creating a euphoric state is becoming increasingly common. Methods used for this abuse, including “sniffing” (directly inhaling vapours), “huffing” (placing a hydrocarbon-saturated rag over the mouth and nose and then inhaling), or “bagging” (inhaling via a plastic bag filled with hydrocarbon vapours) (Barnes, 1985). Petrol sniffing is a major problem in Aboriginal communities across four Australian states. It destroys health and families. It has claimed over 100 Indigenous lives from 1981 to 2003 across Australia. The practice was first observed in 1951, and is believed to have been introduced by US servicemen stationed in the nation’s top end during World War II (MacLean, 2007).

HOW TO RECEIVE PROJECT MATERICAL(S)

After paying the appropriate amount (#5,000) into our bank Account below, send the following information to

08068231953 or 08168759420

(1) Your project topics

(2) Email Address

(3) Payment Name

(4) Teller Number

We will send your material(s) after we receive bank alert

BANK ACCOUNTS

Account Name: AMUTAH DANIEL CHUKWUDI

Account Number: 0046579864

Bank: GTBank.