WeCreativez WhatsApp Support
Welcome! My name is Damaris. I am online and ready to help you via WhatsApp chat. Let me know if you need my assistance.
The Complete Material for this Topic is Available for Download.

Download this complete Project material titled; TKinetic Studies Of The Reaction Of Formaldehyde With Plasma Albumin In Water And Ethanol – Water Mixtures with abstract, chapters 1-5, references, and questionnaire. Preview Abstract or chapter one below

  • Format: PDF and MS Word (DOC)
  • pages = 65

 3,000

100% Money-Back Guarantee

Contact us - 08060082010, 08107932631, 08157509410, Edustoreng@gmail.com

ABSTRACT

Formaldehyde is a simple member of the aldehyde family and one of the simplest organic molecules. It is known to be ubitiquious in nature in foods, domestic air, cigarette smoke etc and has been found to be toxic over certain doses. The chances of harmful effects are reported to increase under room temperature because of its volatility and have also been reported as occupational and environmental toxicant. In spite of these claims, there is paucity of knowledge of the reaction kinetics and mechanisms of this organic molecule with carrier proteins, common non-foods and food materials in the literature. In this study an investigation of formaldehyde levels in some selected foods, non-foods, cosmetics, human milk and urine samples based on spectrophotometric method of Hantzsch by formation of a derivative for separation was carried out using acetylacetone and acetoacetanilide reagents. A 3,5-diaphenyl-1,4-dihydropyridine adduct was also prepared from acetoacetanilide, formaldehyde and plasma albumin and was characterized by measuring its melting point, IR analysis, Uv spectra as well as the antimicrobial activity. The stoichiometry and the kinetic measurements were carried out at constant conditions with the concentrations of the formaldehyde in excess of 0.27 x10-1 mol dm-3 while that of plasma albumin was kept constant at 0.51 x10-4 mol dm-3. The effect of dielectric constant, added ions , pH and ionic strength and acidity constant (pKa) on the rate of reaction were all monitored by varying their concentrations while keeping the concentration of the aqueous formaldehyde and all the other conditions constant The kinetic of the reaction of plasma albumin with formaldehyde in water and ethanol-water mixtures was also investigated using bronsted-type plot model. Both the acetylacetone and acetoacetanilide reagents methods found formaldehyde levels in the studied samples at a non significant amount with a (p > 0.0001) at 95% degree of confidence. Also the
viii
prepared 3,5-diaphenyl-1, 4-dihydropyridine adduct was found to have a melting temperature range of 179.5 – 180.4o indicating that it was a labile schiff base and a protein with glycol protein (lectin) properties. The stoichiometric data showed a mole ratio of 1:2 for formaldehyde to plasma albumin and the reaction was found to be pseudo-first order with respect to formaldehyde and overall second order at constant conditions in both systems. The rate of the reaction in water medium and ethanol water-mixtures was found to increase with increased pH, ionic strength and the added ions. The results of the activation parameters in water and ethanol-water mixtures showed a large negative entropy, small positive values of activation energy, negative values of enthalpies of reaction and a small positive Gibbs free energies. This shows that the reaction was not spontaneous and thermodynamically hindered or unfavourable but can take place by a collision or activated complex mechanism. The values of rate constants of the reaction between plasma albumin and formaldehyde were found to decrease with the increase in the concentration of ethanol and the reaction. The corresponding values of the Brønsted-type plots proportionality constants (β) for the reaction in water solution and ethanol-water mixtures were found to be β = 0.059 and 0.0021 respectively. The second order rate constant values of 0.099 (±0.034) dm3 mol-1 s-1 in water and 0.06 (±0.028) dm3 mol-1s-1 ethanol-water mixtures respectively were obtained. Also appropriate mechanistic proposals were made for the reaction in water and ethanol-water systems. The proposals showed that one mole of formaldehyde binds two moles of plasma albumin forming Schiff bases slowly and reaction was found to operate by binding/ associative mechanism
ix

 

 

TABLE OF CONTENTS

Cover page i Fly leaf Title page ii Declaration iii Certification iv Acknowledgements v Dedication vi Abstract vii Table of Contents ix List of Tables xvii List of Figures xxi Abbreviations xxv List of Appendices xxvii List of Publications from this thesis xxix CHAPTER ONE 1.0 INTRODUCTION 1 1.2 The Research Problem 8 1.3 Justification of the Research 10
1.4 The Research Hypotheses 12
1.5 Aim and Objectives of the Study 13 CHAPTER TWO 2.0 LITERATURE REVIEW 14 2.1 Formaldehyde 14 2.2 Synthesis and Industrial Production of Formaldehyde 14
x
2.3 Sources of Formaldehyde 15 2.4 Uses of Formaldehyde 16 2.5 Toxicity of Formaldehyde 17 2.5.1 Case reports and clinical studies on formaldehyde 17 2.5.2 Formaldehyde as a risk factor for Cancer 18 2.5.3 Laws and actions of some countries on the use of formaldehyde 20 2.6 Methods used for the Determination of Formaldehyde 21 2.7 Blood Plasma Albumin 22 2.8 Plasma Albumin Chemistry 24 2.8.1 Structure of serum albumin 25 2.9 Isolation and Purification of Plasma Albumin from Human Blood Samples using Organic Solvents 27 2.10 Formaldehyde Stabilizers 28 2.11 Reaction between Proteins and Formaldehyde 29 2.12 Reaction of Proteins Tau with Formaldehyde 33 2.13 Effect of Solvent Medium on Reactions 35 2.14 Rate Mechanisms in Binary Mixtures 37 2.15 The Effect of Dielectric Constant on the Reaction Rate 43 2.16 pK Values of Some Amino Acids Reacting Groups 44 2.17 Determination of a Ligand to Ligand or Ligand to Metal Ratio 48 2.17.1 The method of continuous variation (Job’s method) 48 2.17.2 Mole ratio method 49 2.18 Formaldehyde-Plasma Albumin Adduct as a Glycoprotein (Lectin)
xi
CHAPTER THREE 3.0 MATERIALS AND METHODS 50 3.1 Reagents 50 3.2 Methods 51 3. 2.1 Sampling 51 3.2.2 Extraction of formaldehyde from human urine, milk, foods, cosmetics and non – food materials 55 3.3 Isolation and Fractionation of Plasma Albumin from Human Whole Blood using Ethanol as Organic Solvent 54 3.3.1 Removal of haemoglobin from the blood samples 54 3.3.2 Removal of α- and ß- globulin 55 3.3.3 Precipitation of pure albumin protein 55 3.3.4 Characterization of isolated plasma albumin 56 3.3.5 Protein analysis by Kjeldahl method 56 3.3.6 Solubility test 57 3.3.7 Denaturation tests 57 3.3.8 Optical rotation measurement of the albumin samples 58 3.4 Synthesis and Characterization of 1,4-Dihydropyridine from Formaldehyde, Acetoacetanilide and Plasma Albumin 58 3.4.1 Fourier transform infrared spectroscopy 59 3.4.2 Melting point analysis 59 3.4.3 Antimicrobial screening test 61 3.5 Preparation of Samples and Reagents 61 3.5.1 Distillation of paraformaldehyde to obtain pure formaldehyde solution 62 3.5.2 Standardization of the purified formaldehyde solution by iodometric method 62
3.5.3 Preparation of formaldehyde working solutions 63
xii
3.5.4 Preparation of plasma albumin working solutions 63 3.5.5 Preparation of percentage ethanol-water mixtures 64 3.5.6 Preparation of percentage ethanol–water mixtures of varying dielectric constants 64 3.5.7 Preparation of hemiacetals from ethanol- water mixtures and formaldehyde concentrations 65 3.5.8 Preparation of acetylacetone – ammonium citrate buffer solution 65 3.5.9 Preparation of ammonium citrate 65 3.5.10 Preparation of aqueous hydrochloric acid solutions 66 3 .5.11 Sodium hydroxide solutions 66 3.5.12 Preparation of sodium chloride solutions 66 3.5.13 Preparation of potassium iodate 66 3.5.14 Sodium thiosulphate 67 3.5.15 Starch indicator solution 67 3.5.16 Iodine solution 67 3.5.17 Acetic acid (10 moldm-3) 68 3.6 Preparation of Formaldehyde Calibration Curve and Determination of Formaldehyde Levels in the Samples using Acetylacetone Reagent 68 3.6.1 Preparation of formaldehyde calibration curve and determination of formaldehyde levels in the samples using acetoacetanilide reagent 69 3.7 Kinetics of Reactions of Plasma Albumin and Formaldehyde in Water Medium 69 3.7.1 Determination of wavelength of maximum absorption (λmax) of formaldehyde and plasma albumin 70 3.8 Stoichiometry of the Reaction of Plasma Albumin and Formaldehyde 70 3.8.1 Photometric titration 71 3.8.2 The Job‘s method 71
xiii
3.8.3 The mole ratio method 72 3.8.4 Determination of rate and order of the reactions of plasma albumin with formaldehyde in water solution 72 3.9 Reactants Concentrations Dependence Studies 73 3.9.1 The effect of formaldehyde concentration 73 3.9.2 The effect of plasma albumin concentration 74 3.10 Effect of Hydrogen Ion Concentration on the Reaction Rate 74 3.11 Temperature Dependence Studies 75 3. 12 Effect of Ionic Strength on the Reaction Medium 76 3.13 Effect of Added Ions on the Reaction Rate 76 3.14 The Effect of Dielectric Constants and at Different Temperatures on the Reaction Rate 76 3.15 Spectrophotometric Determinations of the pKa of Plasma Albumin 77 CHAPTER FOUR 4.0 RESULTS 79 4.1 Analysis of Formaldehyde Levels in Human Urine, Milk and Food-Related Samples 79 4.2 Analysis of Formaldehyde Levels in Cosmetics and Non – Food Materials 79 4.3 Characterization of Isolated Plasma Albumin and Prepared Adduct 80 4.4 Ultra Violet Light Spectra Analysis of 1, 4 Dihydropyridine 90 4.5 Melting Point, Antimicrobial Susceptibility and Infrared Spectroscopy Analyses of 1, 4 Dihydropyridine Adduct 90 4.6 Kinetic Studies 91 4.6.1 Wavelengths of maximum absorption (λmax) of formaldehyde, plasma albumin and formaldehyde – plasma albumin mixture 100
xiv
4.6.2 Stoichiometry 100 4.6.3 Order of reaction with respect to reactants in water medium 100 4.6.4 Order of reaction with respect to reactants in ethanol-water- mixtures 105 4.6.5 The effect of varying plasma albumin concentrations on the reaction rate of plasma Albumin and formaldehyde in water 110 4.6.6 Ionic strength studies on the reaction of plasma Albumin with formaldehyde in water 110 4.6.7 Ionic strength studies on the reaction between plasma albumin and formaldehyde in 15% (Ethanol-water) mixture 110 4.6.8 Temperature dependence studies in water system 117 4.6.9 Temperature dependence studies in 15% and 20 % (ethanol- water) system 117 4. 6.10 pH dependence studies in water system 130 4.6. 11 Effect of added anions on the reaction rate 130 4.6.12 Kinetic study of effect of dielectric constants on the reaction of formaldehyde and plasma albumin in ethanol-water mixtures 140 4.6.13 Results of the kinetic study of reactions of plasma albumin with formaldehyde in ethanol – water mixtures and water using Bronsted-type plot 147 CHAPTER FIVE 5.0 DISCUSSIONS 155 5.1 Analysis of Formaldehyde Levels in Human Urine, Milk and Food- Related Samples 155 5.2 Analysis of Formaldehyde Levels in Cosmetics and Non-Food
xv
Materials 155 5.3 Characterization of Isolated Plasma Albumin and Prepared Adduct 157 5.4 Ultra Violet Spectra, Melting Point, Antimicrobial Susceptibility and Infrared Spectroscopy Analyses of 1, 4- Dihydropyridine Adduct 158 5.5 Kinetic Studies 159 5.5.1 Wavelengths of maximum absorption (λmax) of formaldehyde, plasma albumin and formaldehyde- plasma albumin adduct 159 5.5.2 Stoichiometry 159 5.5.3 Determination of order with respect to reactants in water and ethanol – water mixtures 160 5.5.4 Ionic strength studies on the reaction between plasma albumin and formaldehyde in water and 15% ethanol-water mixture 164 5.5.5 Temperature dependence studies in water and ethanol-water mixtures 166 5.5.6 The effect of hydrogen ion concentration on the reaction rate 167 5.5.7 Effect of added anions and cations on the reaction rate 168
5.5.8 Biochemical reaction mechanism of formaldehyde – plasma
albumin adduct 169
5.5.9 Kinetic study of reactions of plasma albumin with formaldehyde in ethanol – water mixtures and water solution using Bronsted-type plot 171 5.5.10 Mechanistic proposals of the rate of reaction of formaldehyde with plasma albumin in water 173 5. 5.11 Mechanistic proposal of the rate of reaction of formaldehyde with plasma albumin in ethanol – water medium 176 5.5.12 Mechanistic proposals for ethanol-water mixtures based on SESMORTAC model 179
xvi
5.5.13 Mechanistic proposal based on bronsted-plot in water solution and ethanol – water-mixtures 181 CHAPTER SIX 6.0 SUMMARY, CONCLUSION AND RECOMMENDATIONS 182 6.1 Summary 182 6.2 Conclusions 187 6.3 Recommendations 188 6.4 References 189

 

Project Topics

 

CHAPTER ONE

 

1.0 INTRODUCTION
Formaldehyde is a colourless, flammable gas that has a distinct, pungent smell. It is a simple substance containing oxygen, hydrogen and carbon and the simplest member of compounds called aldehyde and a natural part of our atmosphere (WHO, 1989; IARC, 1995). Formaldehyde is a known gas at room temperature with a chemical formula CH2O and it is found to be readily soluble in water. It is also reported to be widely present in aquatic and air environments. Relatively, it is considered to be an important organic compound in industrial synthesis as well as an indoor air contaminant (Khoderet al., 2002). It is reported widely to be present in aquatic and air environments (CEPA, 2004). Formaldehyde is also known as methanal, methyleneoxide, oxymythelene, methyl aldehyde, formalin, morbic acid, oxo methane and embalming fluid and is reported to occur naturally in foods (FDA 1998). There are reported levels of natural formaldehyde in raw foods, ranging from 1 mg/kg up to 90 mg/kg and accidental contamination of food may occur through fumigation, the use of formaldehyde as a preservative, or through cooking (WHO,1989). Tobacco smoke as well as urea-formaldehyde foam insulation and formaldehyde-containing disinfectants are all known sources of indoor formaldehyde (CEPA, 2004). The major anthropogenic sources affecting humans are therefore reported to be from indoor environment (CEPA, 2004). Products containing formaldehyde, such as resins, glues, insulating materials, chipboard, plywood and fabrics, are common. (WHO, 2001).
The most commonly known commercially available forms are 30-40% aqueous solution (formalin), parasite-S, paracide-F and Formalin-F. These are usually used as formaldehyde releasing agents for the preservation of fish (FDA, 1998). Formaldehyde is known to be naturally produced in the human body in small amounts (ATSDR, 1999). It has been reported to occur in blood, different organs, fruits, seeds, micro-organisms,
2
plants and animals and is emitted as a by-product of certain vegetables, such as cabbage and Brussels sprouts when they are cooked (Goldernberg, 1994). It is also produced by high energy irradiation of carbohydrates or proteins and by autooxidation of unsaturated fat and fatty acids. It is known to be one of the volatile compounds formed in the early stages of the decomposition of plant residues in the soil (Goldernberg, 1994). Industrially, formaldehyde is used in the production of cosmetics and sugar, well drilling fluids, in agriculture as a preservative for grains and seed dressing. Other areas of its use include, the rubber industry, the production of latex, leather tanning, wood preservation and photographic film production in hospitals and laboratories to preserve corpse and tissue specimens (US – EPA, 1999). Formaldehyde dissolves easily in water but does not last long enough in it. However, during this period aquatic life in water could come in contact with it before it is oxidized. Exposure to formaldehyde is reported to cause irritation of eyes, nose, throat and skin at low levels in humans while it is known to be carcinogenic at high levels (ATSDR, 2011). This organic substance has been found in at least 26 of the 1,467 of the United States national priorities list sites as identified by their Environmental Protection Agency. The Occupational Safety and Health Administration (OSHA) of the United States of America has set a permissible exposure limit for formaldehyde at 0.75mg/L for an 8hr working day or 40 hrs working week and the United States National Institute for Occupational Safety and Health (NIOSH) recommended exposure limit of 0.01mg/L (NIOSH, 1997). Families can reduce the risk of exposure to formaldehyde vapours by opening windows, the use of fans to bring in fresh air indoors, removing formaldehyde sources in the home, seal of unfinished manufactured wood surfaces and washing of new permanent press clothing before wearing can help lower exposure (ATSDR, 1999).
3
Microbial contamination of household products, especially those used around the eyes and on the skin, can cause significant problems and preservatives can help to prevent such problems of contamination by its anti microbial activity (Moennich et al., 2009).
A report from US Department of Health and Human Services (US, DHHS) showed that Formaldehyde and its releasing agents are widely used as preservatives in foods and most household products like, cosmetics, paint, cloth, baby underwear and toilet tissues, medicinal and industrial products in aqueous forms worldwide and has been reported to be first used as a biological preservative more than a century ago (US, DHHS, 2008; CIO, 2008). Formaldehyde-releasing preservatives are reported to be ingredients that are highly efficient in helping to ensure the safety of products by protecting them against contamination by micro organisms during storage and during continued use by consumers. They have the ability to release formaldehyde in very small amounts over time. The use of formaldehyde-releasing preservatives ensures that the actual level of free formaldehyde in the products is always very low but at the same time sufficient to ensure absence of microbial growth. If pure formaldehyde was used, addition of enough formaldehyde at the beginning to ensure preservation over the whole lifetime of a product would be necessary, because formaldehyde is slowly used up over time (CIO, 2008). Formalin which contains 37% w/v formaldehyde has been used as a therapeutant to control echo parasites and aquatic fungi disease event occurring at fish culture facilities. Commonly the fish are dipped in formaldehyde for this purpose. Its residue in food for human consumption are proscribed because it is a possible carcinogen (FDA, 2006). It is also known as a common solvent for tissue fixation and for corpse preservation as well as a bacteriostatic agent in some foods, such as cheese. Fruits and vegetables are found to typically contain 3–60 mg/kg, milk and milk products about 1
4
mg/kg, meat and fish 6–20 mg/kg and shellfish 1–100 mg/kg (Estani, 1992). Formaldehyde is a known toxin to humans. Its residue in food for human consumption are proscribed because of it‘s a possible carcinogen (Kiernan, 2000). The International Agency for Research on Cancer (IARC) and the World Health Organization (WHO) therefore classified formaldehyde as a ―probable human carcinogen‖ for a rare form of nasal cancer (IARC, 2004). The U.S. Environmental Protection Agency (US-EPA) has similarly classified formaldehyde as a probable human carcinogen under conditions of unusually high or prolonged exposure. Formaldehyde is not typically added directly to cosmetics and personal care products, other than as a component of some nail hardening products. Regarding inhalation of large amounts of formaldehyde, the IARC considered new studies again in 1995 and maintained the rating at 2A (probably carcinogenic to humans) (IARC, 1995). IARC also recommended a reclassification of formaldehyde as a known human carcinogen based on provision of sufficient evidence to establish that exposure to formaldehyde causes nasopharyngeal cancer in humans under certain circumstances. (IARC, 2004). The IARC also found strong evidence of a link between formaldehyde and leukemia, though the evidence was not sufficient to establish a causal relationship. Similarly, travel trailers and mobile homes provided by the United States Federal Emergency Management Agency (FEMA) between 2006-2007 for the displaced Gulf coast residents of Hurricane Katrina and Hurricane Rita victims were found to have average levels of 77gL-1 (FEMA, 2007). Exposure to these levels were reported to cause breathing difficulties, nose bleeds and persistent headaches to the residents. Long term exposure to levels in this range was reported to be linked with an increased risk of cancer and of respiratory illness. The levels detected were higher than those expected in indoor air where levels are commonly in the range of 10-20 gL-1.and were higher than those for Agency for Toxic Substances Disease Registry division of
5
the United States Canters for Disease Control and Prevention minimal risk level (ASTDR, 2006). Other reported exposures include cosmetic products containing formaldehyde. Formalin and/or Paraformaldehyde is found to come into contact with hair (e.g. shampoos and hair prep arations), skin (deodorants, bath products, skin preparations and lotions), eyes (mascara and eye make-up), oral mucosa (mouthwashes and breath fresheners), vaginal mucosa (vaginal deodorants) and nails (cuticle softeners and nail creams and lotions (Shimadzu,1973). Exposure from most of these sources was reported to be localized, although some formaldehyde was known to be available for inhalation (e.g. from shaving creams). Systemic absorption, including penetration into the circulatory system, is estimated to be negligible. There have been reports of newborn infants being exposed to formaldehyde containing disinfectants in incubators (Piletta-Zenin et al., 2007).
The role of formaldehyde in mis-folding of neuronal tau protein, and the toxicity of formaldehyde-induced tau aggregates on human neuroblastoma cells has also been investigated (Chun et al. 2007). Formaldehyde, as a cross linking agent was found to react with thiol and amino groups, leading to protein polymerization (Yu et al., 2006). Furthermore, ethanol and methanol ingestion have been proved to be important public health concern because of the selective actions of their toxic metabolites namely formaldehyde and formic acid, on the retina, the optic nerves and the central nervous system (Yu, 2001). In the liver and retina, majority of methanol was found to be converted to formaldehyde, by alcohol dehydrogenase. Formaldehyde was then converted to formate by aldehyde dehydrogenase and other enzymes (Jacobsen and McMartin, 1997). In semicarbazide-sensitive amine oxidase (SSAO)-mediated pathogenesis of Alzheimer’s disease, formaldehyde was shown to interact with β-
6
amyloids to produce irreversibly cross-linked neurotoxic amyloid-like complexes (Yu et al., 2006; Yu; 2001; Gubisne-Haberle et al., 2004). Other studies have also shown that formaldehyde exposure leads to formation of DNA/protein crosslink, a major mechanism of DNA damage. The DNA/protein cross links have been used as a measure of dose in drug delivery (WHO, 1989; Heck and Casanova, 1999). Formaldehyde is also known to be one of the products of incomplete combustion which is released to atmosphere (Yuan, 1985). In metropolitan cities, formaldehyde is reported to be the predominant aldehyde emitted by automobiles especially when alcohol-based fuels are used and also as a secondary product of photochemical reactions of volatile organic compound (Lei et al., 2010).The main sources for the formaldehyde pollution in air are reported to include painting, coating material and cigarette smoking (US, EPA, 2012). However, formaldehyde is known to be toxic to humans and other animals at high concentrations and particularly toxic to humans at levels above 0.08% w/v of formaldehyde in human blood and irritating to the respiratory tract, eye and skin (Chun et al., 2007). Inhalation of a large amount of formaldehyde is reported to cause severe nasal tumours and irritation of the upper respiratory tract in rats, mice and humans. Data from human exposures indicated that high concentrations of formaldehyde may lead to pulmonary edema (IARC, 2007).
The daily intake of formaldehyde has been reported to be difficult to evaluate, but a rough estimate from the available data is found in the range of 1.5 –14 mg/day for an average adult, most of it in a bound and unavailable form WHO, 1989, WHO, 2001). A sufficient amount of formaldehyde reaching target cells, and the saturation of formaldehyde metabolism to formate is reported to increase the covalent binding of formaldehyde to deoxyribonucleic acid (DNA). The workers were of the view that the
7
carcinogenicity of formaldehyde could result from its ability to induce DNA-protein cross-links or hydroxymethyl adducts in DNA (Fennel,1994). Formaldehyde has also been described as one of the chemical mediators of apoptosis, that is programmed cell death (Abubakar et al., 2009). The presence of formaldehyde residue in the food meant for human consumption are therefore proscribed because of possible carcinogenicity (Jung et al., 2001). Moreover, the formaldehyde generated in situ from some N-methylated compounds have been reported to induce apoptosis or retardation of cell proliferation to tumorous cells known as cancer (Korpan et al., 2000). ‗ Recent studies have shown that neuro degeneration is closely related to misfolding and aggregation of neuronal tau. Neuronal tau is an important protein in promoting and stabilizing the microtubule system involved in cellular transport and neuronal morphogenesis. The significant protein tau aggregation induced by formaldehyde and the severe toxicity of the aggregated tau to neural cells may suggest that toxicity of methanol and formaldehyde ingestion is related to tau misfolding and aggregation. Both formaldehyde and acetaldehyde can go through the blood-brain barrier and cause some lesions to CNS, especially our visual system. Formaldehyde is reported to penetrate through the blood-brain barrier and could cause some lesions to central nervous system, especially our visual system (Scherbakova et al., 1986). It has also been established that high levels of formaldehyde react with proteins and nitrogen atoms in the environment to form reversible and irreversible adducts in vitro and in vivo and clinically, the lethal dose of formaldehyde for human beings is found to be about 0.08% w/v in the circulation (Erkrath et al.,1981). 1.2 The Research Problem
8
Formaldehyde levels higher than those obtainable in typical indoor exposure levels of formaldehyde have been found in travel trailers and mobile homes used as emergency housing for displaced residents of Gulf Coast Region, United State of America by Hurricane Katrina and Hurricane Rita (FEMA, 2007; Harris et al., 1981). Long-term exposure to Formaldehyde has been found to cause irreversible neurotoxicity implicated in cancer of the central nervous system (brain astrocytoma) (Kilburn, 1994, Stroup, et al., 1986). In addition, inhalation of formaldehyde has been shown to cause behavioral and memory disorders in rats and has been classified as a probable neurotoxic agent (Pitten et al., 2000). It has also been reported that inhaled formaldehyde gas has negative effects on the central nervous system, and these effects may appear acutely in the form of headaches, malaise, sleeping disorders, fatigue, anorexia and dizziness. Formaldehyde has been reported to have the potential of significantly mis-folding DNA and native soluble proteins into attendant amyloid fibrillation and insoluble fibrils comprising of cross- ß-sheets (Chun et al., 2005; 2007). Formaldehyde has also been found to have the potential of oxidizing protein-adducts with the attendant increase in acid levels in the blood even if it remains in the human body for only a short time (Sipe et al.,2005). Formaldehyde is reported to be clinically toxic to humans at a concentration of 0.08 % w/v in the body (Erkrath, 1981). It has also been described as one of the chemical mediators that cause apoptosis (Abubakar et al., 2009). There are evidences to suggest that under physiological conditions glucose reacts with formaldehyde and other related oxidizing agents non-enzymatically with a wide variety of proteins to form glycated products. Non-enzymatic glycation of proteins which has been shown to be a potential problem during their storage in the food and biotech industries (Davis et al., 2001) and is also implicated in human diseases such as diabetes (Al-Abed et al., 1999; Stitt, 2001) is accelerated in the presence of formaldehyde.
9
Rajbar (1968) reported that human blood proteins like hemoglobin and serum albumin undergo a slow non enzymatic glycation with oxidizing agents such as formaldehyde, mainly by forming schiff bases between ɑ- amino groups of lysine and some times arginine and glucose molecules in the blood. Elevated glycol albumin has been reported in diabetes mellitus (Iberg and Fluckiger, 1986). Glycation is found to result in the formation of advanced glycosylation end products (AGE) leading to abnormal biological effects. AGEs are reported to be antigenic and represent many of the important neo antigens found in cooked and stored foods and also found to be channel blockers for calcium, magnesium, etc (Vazquez et al., 2008). Because of the toxicities of formaldehyde to humans, quest for information about how this substance causes specific disease conditions through its reactions with biomolecules in the human body has become a necessity. To date, there are a few scientific studies reporting the detailed mechanisms of the reaction of formaldehyde with bio-molecules, especially proteins. In fact, Chun et al., (2007) were the only ones known to have reported the kinetics and mechanisms of the reaction of formaldehyde with a human protein, protein tau using a sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) method.Their study found that protein tau was crosslinked and aggregated by formaldehyde via a second order mechanism thus indicating that formaldehyde is neurotoxic. Formaldehyde in the human body is distributed by blood and its possible reaction with plasma albumin is worth exploring. 1.3 Justification of the Research
There has been an increase in the reported negative influences of formaldehyde to nature and humans which has necessitated that the routine analysis of formaldehyde in the environmental niches like water, fish, cosmetics, foods and non- food materials etc.
10
should be urgently carried out (Uzairu et al., 2009). The role of formaldehyde in mis-folding of neuronal tau protein (Chun et al,2005) and the toxicity of formaldehyde-induced tau aggregates on human neuroblastoma cells has also been reported (Chun et al. 2007). Formaldehyde is reported to be a cross linking agent and is known to react with thiol and amino groups, leading to protein polymerization (Yu, 2001; Yu et al., 2006). Furthermore, ethanol and methanol ingestion has been proved to be an important public health concern because of the selective actions of their toxic metabolites, which include formaldehyde and formic acid, on the retina, the optic nerves and the central nervous system (Yu, 2001). In semicarbazide-sensitive amine oxidase (SSAO)-mediated pathogenesis of Alzheimer’s disease, formaldehyde is shown to interact with β-amyloids to produce irreversibly cross-linked neurotoxic amyloid-like complexes (Yu, 2001; Yu et al., 2006; Gubisne-Haberle et al., 2004). It is therefore necessary to not only assess formaldehyde levels in foods, non-food products and body fluids but to kinetically investigate the interations of formaldehyde with other proteins such as plasma albumin in order to evaluate the health risks associated with its trace presence in the blood stream where plasma albumin protein is most abundant. In spite of the above known and documented toxicity of formaldehyde, there are only a few studies exploring its levels in some common foods, non-foods materials and some body fluids and the detail mechanisms of its interactions with the biomolecules in the literature. Furthermore most of the information available on formaldehyde reaction with proteins and biological tissues are only based on fixation reactions from tanning and embalmment industries where high concentrations and volumes of formaldehyde are applied to tissue sections at frozen conditions (Bedino, 2003). In fact to the best of our knowledge, there is no detailed study on the kinetics andmechanism of the reaction
11
between trace levels of formaldehyde with plasma albumin under normal and varying physiological conditions in water solution and binary ethanol-water mixtures or media. Formaldehyde has also been clinically reported to be toxic to humans at a concentration of 0.08%w/v in the body (Erkrath, 1981) and has also been described as one of the chemical mediators that cause apoptosis (Abubakar et al.,2009). There are evidences to suggest that under physiological conditions glucose reacts with formaldehyde and other related oxidizing agents non-enzymatically with a wide variety of proteins to form glycated products. Non-enzymatic glycation of proteins which has been shown to be a potential problem during their storage in the food and biotech industries (Davis et al., 2001) and is also implicated in human diseases such as diabetes (Al-Abed et al., 1999; Stitt, 2001). The glycation reaction is found to be accelerated in the presence of oxidants such as reactive oxygen intermediates, free radicals, formaldehyde etc. Other studies have also shown that Formaldehyde exposure leads to formation of DNA/protein crosslink, a major mechanism of DNA damage. The DNA/protein cross links have been used as a measure of dose in drug delivery (WHO, 1989; Heck and Casanova, 1999).
The significance of the present study therefore is that an investigation of the kinetics and mechanism of the reaction of plasma albmin with formaldehyde in water solution and ethanol – water binary mixtures as well as an assessment of the levels of the latter in some consumable foods and non-food materials and body fluids will provide information on the possible sources of exposure to this toxin to humans. The study will also expose the health risk factors associated with its presence in the blood stream even at trace levels. Such information is expected to guide individuals, public health workers and health care providers as well as policy makers on health on health issues of the
12
dangers associated with elevated levels of formaldehyde and its reaction pathways when exposed to it in the environment. 1.4 The Research Hypotheses This study was based on the following two hypothes (i) Null hypothesis, H: This states that formaldehyde is absent in commonly used food, non- food materials and human body fluids and that formaldehyde does not in anyway cross link native soluble proteins like plasma albumin hence does not constitute a human risk factor. (ii) The alternative hypothesis, H1: This states that elevated values of formaldehyde are present in commonly used foods, nonfood materials and human body fluids and that formaldehyde cross links native soluble proteins like plasma albumin which may cause health hazards in humans.To verify which of the two hypotheses above holds, some selected foods, non-food products, human milk and urine samples were analyzed for formaldehyde. Also, the reaction of formaldehyde with protein albumin in water and various ethanol- water mixtures was studied kinetically under various conditions similar to human blood serum in order to explore the health risks associated with the presence of formaldehyde levels in the human blood. 1.5 Aim and Objectives of the Study The aim of this investigation was to measure formaldehyde levels in some foods, Non food materials and body fluids as well as study the kinetics and mechanisms of the reaction of formaldehyde with plasma albumin in water and ethanol-water mixtures under various conditions similar to human blood. This was hoped to be achieved through the following objectives:
13
(i). To detect and measure amounts of formaldehyde in commonly used foods,non-food materials some body fluids and using acetylacetone and Acetoacetanilide reagents. (ii). To isolate, purify, fractionate and characterize plasma albumin from whole blood samples using ethanol a common organic solvent with the view to studying its reaction with formaldehyde. (iii).To investigate the kinetics of the reaction of formaldehyde with plasma albumin under normal and physiological conditions involving pH, temperature, reactants concentrations and ionic strength in water and (0 – 25%) ethanol – water mixtures. (iv). To propose plausible mechanism(s) for the reactions of formaldehyde and plasma albumin using various kinetic models like Bronsted – plot and rate equations.

 

GET THE COMPLETE PROJECT»
Do you need help? Talk to us right now: (+234) 08060082010, 08107932631, 08157509410 (Call/WhatsApp). Email: edustoreng@gmail.com

DISCLAIMER:This is a complete project material for students and should NOT be copied verbatim, which is Plagiarism. Only use as a guide in developing your original research work. Call/Whatsapp us if you need our help: (+234) 08060082010, 08107932631, 08157509410. Email: Edustoreng@gmail.com