Effects of Atmospheric Aerosols on Human Health

Effects of Atmospheric Aerosols on Human Health Abstract: A highly Sensitive (LOD; 0.04-0.4 ng/ml) method is developed for detection and quantification of acidic compounds (C3 -C10) containing mono and dicarboxylic acids on GC-MS. These compounds (C3 -C10) existed in trace amount, as secondary organic aerosols i.e. important constituents of Aerosols. Membrane extraction technique was utilized for selective enrichment (1-4300 times) of target compounds. Good repeatability (RSD% ≠¤ 10%) from selective organic phase (10% TOPO in DHE) was achieved with three phase HF-LPME. Aerosols containing samples, after Ultrasonic Assisted extraction were detected and quantified Through GC-MS. Effective derivatization of each target compound was performed with BSTFA reagent. Gas Chromatography, having capillary column and interfaced with mass spectrometry was used for separation, detection and quantification of target compounds. Method Development and Application -hollow fiber Supported liquid membrane extraction of Fatty acids (C3-C10) containing mono and dicarboxylic acids and Detection of aerosols Samples after ultrasonic assisted extraction. 1. Introduction: Impact of Atmospheric aerosols on human health and effect on radioactive stability in Earth’s atmosphere is getting importance now a days and this phenomenon has been well understood. [1]. Atmospheric aerosols can harm respiratory and cardiovascular system of human. Impact of Secondary organic aerosols as biogenic and anthropogenic antecedent is identified (Adams and sinfold, 2002) [1, 17]. Low molecular dicarboxylic acids (C3-C9) are also vital tracers of SOA [2]. Short chain fatty acids are found as secondary organic aerosols which are also supposed to derive from long chain fatty acids [1]. Importance of organic aerosol has been well established now a days and carboxylic acids are of great interest for environmental studies [1]. Several studies and mechanisms were proposed to understand the production of these SOA precursors [1]. Short chain carboxylic acids are found extensively in troposphere [2]. Secondary organic aerosols (SOA) are formed in the atmosphere by gas particles conversions. Organic matter present in aerosol contains more than 90% of troposphere’s aerosols [5, 15]. Dicarboxylic acids found in nature as polymeric compounds such as suberin and cutin [3]. Short chain dicarboxylic acids are found in vegetables [Siddiqui, 1989] and in soil containing micro organisms of durum wheat [4]. Dicarboxylic acids are found in plant oils which have greater interest for cosmetic and pharmaceutical industries [6]. Short chain dicarboxylic acids having aliphatic chain possess strong cyclotoxicity and antineoplastic activities [18]. Many analytical techniques are used to determine the composition of SOA so keeping in view these techniques new method for determination of fatty acids (common in SOA) has been developed. Membrane extraction is used in this method due to its increasing importance for high selectivity and high enrichment factor [24]. Dicarboxylic acids formed of bio oxidation of fatty acids so these are considered as metabolic part of fatty acid [42]. Dicarboxylic acids and their derivatives can be used to make polymers and their condensation with diols in solution produces high molecular weight polyester [39]. Additionally these dicarboxylic acids use less temperature in the reaction for the preparation of polyesters [39]. 1.1. Analytes Description: Properties (physical, chemical, etc.) of Compounds (C3-C10) containing mono and dicarboxylic acids are discussed in section; 1.1.1-1.1.12. These compounds (C3-C10) are the target analytes in this diploma project. These target analytes are extracted through Liquid phase micro extraction and detected by GC-MS system. Fig. 1.1-1.12 represents structures of target analytes (section; 1.1.1-1.1.12). 1.1.1- Adipic Acid Adipic acid is a product of lipid per oxidation. Adipic acid does not undergo hydrolysis in the environment perhaps due to the lack of hydrolysable functional groups (Harris 1990) [5]. 1.1.2- Malonic Acid: Malonic Acid is a metabolite of plants and tissues and Malonyle-CoA [28]. Malonic Acid is an intermediate for preparation of fatty acids from plants and other tissues [7]. Malonic acid is also present in aerosols [8]. Malonic acid is an important constituent of short chain fatty acids [8]. Malonic acid present in beet rots as a Calcium salt [42]. 1.1.3- Succinic Acid: Succinic acid is found in atmosphere as water soluble compound and as a compound of Secondary organic aerosols [29]. Succinic acid is a solid exists as crystals, anciently called spirit of amber. Succinic acid is an important intermediate in citric acid cycle which is very important constituent of living organism [42]. 1.1.4- Glutaric acid: Glutaric acid is found as SOA in aerosols [8]. Glutaric acid is sparingly soluble in water [41], can be used to prepare a plasticizer for polyester [41]. 1.1.5- Pimelic Acid: Pimelic acid is a last dicarboxylic acid relative to carbon number which has IUPAC name. Derivatives of Pimelic acid are used for biosynthesis of amino acid typically lysine [41]. Pimelic acid is produced, when Nitric acid is heated with Oleic acid as a secondary sublimation product which is not crystallized [20]. 1.1.6-Suberic Acid: Suberic acid is produced from suberine [8]. Suberic acid can also be obtained by vigorous reaction condition of natural oil with nitric acid [8]. 1.1.7-Azelaic Acid: Azelaic acid is an important constituent of secondary organic aerosols because it produces short chain fatty acids upon photo oxidation and also because it can be produced during oxidation of unsaturated acid that is found in Oleic acid [11]. 1.1.8- Cis-pinonic Acid: Cis-pinonic acid is also produced in atmosphere by photo oxidation of ÃŽ ±-pinene in the existence of Ozone [30]. 1.1.9- Pinic Acid: Pinic acid is derivative of ÃŽ ±-pinene. Pinic acid can be generated by photo oxidation of ÃŽ ±-pinene with Ozone as given in this chemical reaction; (C10H16 + 5/3 O3 -> C9H14O4 + HCHO). Pinic acid is present in a crystalline form used to prepare plasticizers [30]. 1.1.10- 4-Hydroxybenzoic Acid 4-Hydroxy benzoic acid is exists as crystals. It is used to derive parabens and can be used as antioxidant [41]. 1.1.11-Phthalic Acid: Phthalic acid is an aromatic dicarboxylic acid it is found as white crystalline state in pure form [41]. Phthalic acid is found abundantly in atmosphere and it has toxic properties. Aromatic acids are generally emitted through anthropogenic sources like reminiscent of solvent evaporation and Automobile exhaust [31]. 1.1.12-Syringic Acid. Syringic acid is found as humic substance in environment [40]. 1.2. Detection of Ultrasonic Assisted Extraction samples(UAE): A detection procedure by GC-MS is established with reference standard injections and UAE samples. A theoretical description is given in section 1.2 for â€Å"Ultrasonic assisted extractions†. Unknown real Samples from Aerosols containing mono and dicarboxylic acids (C 3-C 10) are provided after Ultrasonic assisted extraction [34]. 1.2.1- Ultrasonic Assisted Extraction: ‘Ultrasonic’ is derived from ultrasound. Ultrasound refers to a sound that has a higher frequency than a normal human can hear. This technique is used in chemistry in several aspects and due to application in chemistry it is known as Sonochemistry [23]. Ultra Sound is used in sample preparation in analytical chemistry like extraction, filtration, dissolution and sample purification. When Ultrasonic technique is used for assistance in extraction, this assistance in extraction is called â€Å"Ultrasonic assisted extraction† (UAE) [23]. There are many advantages by using UAE because it require less organic solvents ,non destructive, less expensive and less time consuming comparative to other sample preparation techniques like soxhlet [21]. The normal range of ultrasound frequencies used in laboratory ranges from 20 KHz to 40 KHz. Use of UAE is simple. A sample solution inside a vessel in an appropriate solvent can be placed inside ultrasonic bath at desired temperature and sound waves stir the sample [20]. The mechanism of US is as â€Å"when a sound source produces a high frequency waves, sample molecules starts vibrating and shift this vibration to other molecules of sample in a longitudinal direction when gas and liquid is used as a sample, while in solid sample both longitudinal and transverse waves can be produced† [19]. When UAE is utilized it increases speed of mass transport by vibration of mechanical transport from the sample matrix through a process called â€Å"cavitation† [21]. 1.2.2- Theory of Ultrasonic Assisted extraction: There are two theoretical aspects of sonication i.e. physical and chemical aspects in sample preparation. Physical and chemical aspects are described in section (1.2.2.1-1.2.2.2), in order to understand its practical use in analytical chemistry. 1.2.2.1- Physical aspects of UAE: During Ultrasonic assisted extraction, a bubble in a liquid cannot take energy (due to US) and implodes. On the other hand due to Ultra sound in liquid extractions, the cavitational pressure is shifted relatively higher so formation of bubble is difficult [21]. Ultrasonic intensity produces cavitations in a liquid sample during extraction (UAE). Two types of US cavitation is produced known as â€Å"transient cavitation† (produce transient bubble) and â€Å"permanent cavitation† [21]. The life time of transient bubble is so short that no mass transport or diffusion of gas is possible with in a sample [21]. Transient bubble is believed to be produced at US intensity (10 W/cm2) and permanent bubble at intensity (1-3 Watt/cm2). Sonochemical effects are intense inside the bubble because energy (numerous amounts) is produced during bubble eruption and production [21]. 1.2.1.2 Chemical aspects of UAE: When US radiation strikes a water molecule it produces free radicals OH* and H* due to collapsing cavitations’ bubble which exhibits high temperature and pressure inside and also many other radicals can be produced in solution [21]. Radical OH* is believed to be more stable and can begin many new reactions while H* radical is not stable. Second Sonochemical effect is pyrolytic reactions that occur inside bubble and can degrade compounds under analysis [21, 23]. 1.3. Liquid Phase microExtraction(lpme): The application of membrane extractions in analytical chemistry has taken the intentions of analysts during recent time. The goal of utilizing membrane extraction is to achieve high enrichment, selective extraction and environmental friendly procedure [24]. Small quantity of solvent (usually in micro liters) is required comparative to old techniques of extractions (soxlet) [24]. Clean extracts are obtained and after extraction, recovered compounds are shifted to another analytical instrument like Gas chromatography or liquid chromatography directly for further quantitative analysis [24]. 1.3.1 Hollow fiber membrane extraction: Two types of membrane are used in LPME. One membrane is flat sheet porous and second membrane is polypropylene hollow fiber. In this project polypropylene hollow fiber is used as a membrane support in membrane extractions due to limited cost and to reduce carry over problems [24]. 1.3.1.1 HF-LPME Technique: When a hollow fiber is used in LPME, this technique (LPME) is called hollow fiber liquid phase micro extraction (HF- LPME). In HF- LPME technique, a hollow fiber is used containing a thin film of immobilized liquid membrane inside the pores while the fiber is dipped into an aqueous phase containing objective analytes. Target analytes can transport through the membrane into a liquid filled inside the lumen of the fiber, which is termed as accepter solution [22]. Extraction of target analytes (C3-C10) was carried through three phase HF- LPME during whole of the project. Donor solution was contained analytes in aqueous medium, a suitable organic solvent i.e. Dihexyl ether (TOPO mixture) was used in pores of hollow fiber as a stationary liquid membrane support (SLM). Accepter solution was in aqueous medium [22].Target analytes were recovered into accepter phase after evaporation of water. Acetonitrile solvent was added in dried GC vial along with derivatizing reagent. After derivatization these samples were injected into a Gas chromatographic system. 1.3.2 Basic Principle of LPME: Basic principle is same for all LPME techniques (two phase or three phase LPME), the variation is only from accepter region [24]. In three phase liquid phase micro extraction technique (HF- LPME) a donor aqueous solution is filled in a vial or flask containing sample analytes. A short piece of hollow fiber is used and accepter solution is injected inside fiber through a micro syringe after injecting accepter solution one end is closed and other end contains syringe needle. Fiber containing solutions is inserted in an appropriate organic solvent having less polarity (Dihexyl ether) to create a stationary liquid membrane (SLM). Donor solution pH is adjusted such that it can restrain the ionization of target analytes [22]. The process of three phase extraction [22] can be explained as follows in Eq 1.1. Where ‘A’ is a target analyte, ‘K1’, ‘K2’, ‘K3’ and ‘K4’ are first order extraction rate constants. In order to obtain combined distribution coefficient, at equilibrium recovery, Eq. 1.2 is derived [22]. D accepter/sample = C eq accepter / C eq sample = C Org sample/ C eq accepter =ÃŽ ± D .Korg/sample / ÃŽ ± a. Korg/accepter†¦Ã¢â‚¬ ¦Ã¢â‚¬ ¦Ã¢â‚¬ ¦.1.2 In Eq. 1.2, C eq accepter, C eq sample and C Org sample are the concentration of analytes at equilibrium, in accepter phase, in aqueous sample phase and in organic membrane phase respectively.Here Korg/sample, Korg/accepter are the partition ratio’s between Organic phase and sample phase and between accepter phase and organic phase respectively [22]. ÃŽ ± D and ÃŽ ± a are the extractable fraction of total concentration of target analyte in sample and accepter respectively. If conditions are similar between sample and accepter, other than ionization of analytes in sample phase, from Eq. 1.2, equilibrium is independent from partition ratio of stationary liquid membrane in three phase lpme i.e. it depends mainly on ionization of analytes in sample [22]. Extraction efficiency (E) can be calculated from Eq. 1.3[22]. V sample, V accepter and V mem , in Eq. 1.3, are the volume of donor sample phase, aqueous accepter phase and organic immobilized membrane liquid phase respectively. D accepter/sample and D Org/sample are individual distribution coefficients relative to accepter phase to sample phase and Organic phase (SLM) to sample phase respectively [22]. Eq. 1.3 is derived for three phase lpme. It is evident; from the interpretation of Eq. 1.3 that efficiency is mainly controlled by individual distribution coefficients. Individual distribution ratios are directly dependent on partition coefficients, so by increasing the partition ratios efficiency can be improved [22]. Partition coefficients can be improved by properly adjusting the pH of donor and accepter and by using an appropriate organic solvent. Volume of sample and organic phase should also be kept minimum, according to Eq. 1.3 in order to develop efficiency [22]. 1.3.3-Mass transfer in LPME: Enrichment factor (Ee) for three phase LPME is given in Eq. 1.4. Ee = C accepter/C initial = V sample. E / V accepter †¦Ã¢â‚¬ ¦Ã¢â‚¬ ¦Ã¢â‚¬ ¦.. 1.4 In Eq. 1.4, C accepter is the concentration of target analyte, present in final stage inside accepter solution [22]. When an acidic analyte is ionized in aqueous solution, total extractable fraction of analyte (ÃŽ ±) is given in Eq. 1.5 [24]. ÃŽ ± = [AH]/ [A-][AH] = 1/[1+10(pH-pKa)] †¦Ã¢â‚¬ ¦Ã¢â‚¬ ¦.. 1.5 In the context of Eq. 1.3, the overall distribution constant (D) at equilibrium can be rearranged as given in Eq. 1.5 [24]. D = 1+10 s (pH-pKa) . KD /1 + 10 s (pH-pKa). KA †¦Ã¢â‚¬ ¦Ã¢â‚¬ ¦Ã¢â‚¬ ¦.. 1.6 ‘s’ is equal to 1 for acidic analytes (Eq. 1.6). ‘pKa’ is dissociation constant and pH refers to donor or accepter solution(Eq. 1.6) [24]. à ¢Ã‹â€ Ã¢â‚¬  C = ÃŽ ±D .Cs ÃŽ ±a CA.KA/KS †¦Ã¢â‚¬ ¦Ã¢â‚¬ ¦Ã¢â‚¬ ¦ 1.7 Eq. 1.5-1.7 are derived from Henderson-Hasselbalch relation, in this equation ÃŽ ± represents the extractable fraction of analytes [24]. The driving force for the extraction in neutral conditions of three phase LPME is the concentration gradient (à ¢Ã‹â€ Ã¢â‚¬  C) from sample to accepter [12]. The concentration gradient between two phases, between donor and accepter, is described in Eq. 1.7. K represent partition ratio of uncharged analyte between the membrane and aqueous phase. CA and Cs are the concentrations of analytes in accepter and sample phase respectively. 1.3.4 End point for extraction: Three end points are normally considered for extraction [22]. 1. Exhaustive extraction. 2. Kinetic extraction. 3. Equilibrium extraction. 1.3.4.1 Exhaustive extraction: Exhaustive end point is the specific end point (time), when all amount of analytes are exhausted (which can be practically possible) present in donor [22]. In this practical diploma work, Exhaustive end point will be applied in (LPME) extractions. Enrichment factor will increase by growing analyte concentration in accepter by the passage of time, at certain point it reaches a stable value [12]. Mass transfer between organic phase and liquid phase is dependent on concentration gradient [12]. Enrichment factor can be improved by increasing the value of ÃŽ ±D preferably close to unity and decreasing the value of ÃŽ ±A to zero. Such conditions for the ÃŽ ±D and ÃŽ ±A values are called â€Å"infinite sink† conditions, normally required for exhaustive extractions [22]. Situation close to these values can be achieved for acids by selective tuning the pKa values. For example for acidic compound if pH of accepter is adjusted, 3.3 (pH) units above than the pKa of acidic analytes this Di fference set the value of ÃŽ ±A to 0.0005, at this point accepter can capture all analytes. At this set value (ÃŽ ±A), enrichment factor increases linearly with time [12]. Peak time of enrichment factor, when other parameters are constant, can be calculated by comparison of CA maximum. CA maximum (‘CA’ is considered as time dependent) can be obtained by careful calculation of CA maximum values at a certain time, before this value starts to decrease again [12]. 1.3.5 Rate of LPME: Two parameters, govern the rate of extraction (when extraction approaches to equilibrium conditions), are membrane controlled extractions or diffusion controlled extractions [13, 24]. The maximum concentration Ee can be obtained when concentration gradient (à ¢Ã‹â€ Ã¢â‚¬  C) is approaches to zero described in Eq. 1.8 [13, 24]. Ee (max) = (C a / C d) max = ÃŽ ±D/ÃŽ ±A †¦Ã¢â‚¬ ¦Ã¢â‚¬ ¦Ã¢â‚¬ ¦.. 1.8 In membrane controlled extractions, the rate limiting step is the diffusion of target analytes. When analytes pass through the organic phase, the mass transfer (Km) is given in Eq. 1. 9 [13, 16]. Km  µ K.D m /h m †¦Ã¢â‚¬ ¦Ã¢â‚¬ ¦.. 1.9 In Eq. 1.9; K is partition coefficient, Dm is membrane diffusion coefficient and ‘h m ‘ is the thickness of membrane [13, 16]. 1.3.6 Addition of Trioctylphosphine oxide(TOPO): Mass transfer can be improved for acidic analytes by using different concentrations (w/v) of TOPO in organic solvent typically for short chain carboxylic acids. Interaction of TOPO with polar acids in solution takes place efficiently due to hydrogen bonding [16]. 1.3.7 Trapping of Analyte in Three phase lpme[24]: Concentration enrichment of analytes in three phase LPME can be achieved by stable mass transfer through the membrane to accepter phase. Back diffusion of analytes is prevented by trapping of analytes in accepter phase. In order to achieve high enrichment of acidic analytes pH of accepter phase is fixed enough basic so that when acidic analytes reached to the accepter solution becomes charged. Analytes could not be driven back to donor. So this trapping of analytes due to pH adjustment is called ‘’direct trapping’’. For high enrichment purpose, pH of accepter is usually adjusted 3.3 pH units higher than the pKa values of acidic target analytes while extracting from acidic donor. Buffer capacity of accepter should be sufficient such that during extraction protons from acidic donor cannot be neutralized by the concentration gradient between two aqueous phases during three phase lpme [24]. 1.3.8 Selection for Organic phase: Choice of organic solvent has basic importance in method validation because this solvent directly affect partition coefficient. Organic phase solvent should have low solubility in water [22] and low volatility to prevent solvent losses during extraction process [16]. Organic phase should have high distribution coefficient, between donor to organic phase and between organic to accepter phase, to achieve high enrichment. Organic phase should have adequate affinity to the hollow fiber. Organic phase should be immobilized sufficiently to cause efficient trapping of analytes in the pores through polarity matching [22]. Mixture of organic solvents can also be used as mobile phase [16]. In this project organic solvent is either pure DHE or DHE is also mixed with different amount of TOPO (section; 1.3.6) to achieve high stability of organic phase [22, 24]. 1.3.9 Agitation of sample: Extraction kinetics can be improved by agitation. Agitation increases analyte diffusion from donor to accepter. Organic membrane solution (DHE) is very stable inside pores of the membrane. Shaking by a magnetic stirrer helps analyte transfer from donor solution to the accepter solution [17]. When Donor solution containing analytes is stirred at high speed, probability of fresh solution contact with membrane phase is enhanced [9]. In order to enhance mass transfer all membrane extractions in this project are assisted through agitation by a magnetic stirrer. A membrane extraction assembly is shown in Fig. 1.13. 1.3.10Volume of donor and acceptor solutions. Volume of donor and accepter solution is very important because sensitivity can be improved by proper volume adjustment of accepter solution. Volume of accepter solution should be minimum comparative to donor to get better sensitivity [17]. Volume of accepter solution should be enough to be injected, detected and quantified by GC or HPLC. Volume of the accepter solution should be enough to fill lumen of hollow fiber appropriately [17]. 1.3.11 Adjustment of pH. Proper adjustment of pH of donor and accepter is very important because high partition ratio can be obtained in three phase lpme by proper adjustment of donor and accepter solution [17]. According to Eq. 1.7, Efficiency can be improved by increasing concentration gradient which depends mainly on pH. In this project three phase lpme is utilized on acidic analytes (C3-C9) containing carboxylic and hydroxyl groups so in donor solution pH is adjusted slightly lower than the pKa values of analytes to suppress ionization of these analytes [17]. 1.4. Detection and quantification of Analytes: 1.4.1-GC-MS analysis: GC-MS is a powerful detection technique for environmental trace analysis due to its high sensitivity [14]. Aerosols are existed in trace level so their detection requires a sensitive device with low limit of detection. GC-MS suffers less matrix effect and is usually cost effective and highly selective [14]. Analytes are separated according to their charge to mass (m/e) ratio after passing through mass spectrometer. Scan mode is used for identification of each analyte [14]. When gaseous analytes come to mass spectrometer they are converted to their respective molecular ions. Electron ionization in mass spectrometer strikes molecules to fragments [18]. These molecular ions are specific for each analyte and sensitivity and selectivity can be improved through selected ion chromatogram (SIM) [14]. Signal to noise ratio (SNR) is improved through extracted ion chromatogram (XIC) which is selected through SIM mode [14]. SIM mode is used for qualitative and quantitative analysis [14]. Analytes (C3-C10) are polar and non volatile, so these analytes cannot be detected in pure form and separated by using Gas chromatographic column. A derivatization step is necessary to convert Analyte into volatile substances. Derivatization is made to convert carboxylic and hydroxyl functional groups to their respective ester functional group [14]. 1.5. Derivatization: Two derivatization reagents; ‘’N, O-bis(trimethylsilyl) trifluoroacetamide’’ (BSTFA) and ‘’N-(tertbutyldimethylsilyl)-N-methyltrifluoroacetamide’’ (MSTFA) are commonly used for esterification of hydroxyl and carboxylic functional groups before injecting to GC-MS system[14]. Both derivatizing reagents are applied separately and compared prior to GC-MS analysis. 1.5.1- Silylation: Analytes containing carboxylic acids (C3-C10) are introduced to GC-MS after derivatization. Carboxylic acids are converted to their respective trimethyl silyl ester (TMS derivative) by BSTFA. A nucleuphilic attack is taken place by a hetero atom to silicon atom when BSTFA reagent is used as a derivatization reagent [14]. BSTFA is found very efficient to convert hydroxyl groups to respective Silyl ester [18]. Advantage with BSTFA is that its derivative can be injected directly without purification and it can be used for very sensitive detection [18]. BSTFA is non polar and its efficiency can be improved by using BSTFA in Acetonitrile [32]. Chemical structure of BSTFA is shown in Fig [1.14] below. Due to the use of BSTFA reagent in the reaction, a common peak is appeared at m/z= 73, due to [Si(CH3)3]+ molecular ion and at m/z=145 due to [OH=Si(CH3)2]+ molecular ion . when Analytes containing dicarboxylic acids are used for MS analysis, Ion peak is appeared at m/z=147. Ion peak at m/z=147 is appeared due to the [(CH3)2Si=Si(CH3)2]+ molecular ion [18]. 2. Method: 2.1 Membrane extraction: Three phase HF- LPME method is used for extraction. Section 2.1 describes the method for three phase hollow fiber liquid phase micro extraction technique. 2.1.1 Equipment and reagents for Membrane Extraction: Hollow fiber Accurel PP polypropylene (Q3/2) is purchased from Membrana (Wuppertal, Germany). The wall thickness of membrane is 200  µm, Inner diameter 600  µm and pore size is 0.2  µm. Before extraction a 7.5 cm membrane was cut carefully with a fine cutter. After cutting membrane was washed in acetone and dried overnight. A magnetic stirrer, containing multiple stations, model (Ika-werke, Germany) was used for agitation of donor solution. Micro Syringe 50  µl (Agilent, Australia) was used to push accepter solution inside the lumen of membrane and for holding of membrane. pH meter (Mettler Toledo) was used to measure pH for donor and accepter solution. Volumetric flask (Kebo, Germany) was used for extractions (contain donor solution). Milli-Q water was obtained from Millipore gradient system (Millipore, USA). Hydrochloric acid (37%, Fluka) and Sodium hydroxide monohydrate (Fluka) were used to prepare further solutions. Dihexyl ether (97%) was purchased from Sigma Aldrich. TOPO (99%; Aldrich) was used to prepare solutions in DHE (%, w/v). 2.1.2Set up for Membrane Extraction: 2.1.2.1 Donor solution: The pH donor solution was adjusted to 2. All aqueous solutions were prepared in mill Q water and pH was adjusted by adding HCl (0.1M). All Samples were spiked in a dried 100 ml volumetric flask (Germany). This flask was then, filled up to mark with donor solution. Further 5 ml of donor solution was added in same flask in order to dip membrane inside donor solution. Total volume of donor solution was adjusted to 105 ml. A clean magnet was dropped in flask and then, this spiked solution inside the flask was allowed to stir for 30 minutes and at a fixed revolutions/min (800 rpm) of magnetic stirrer. 2.1.2.2 Accepter solution: Accepter solution was prepared in milli Q water and pH 12 was adjusted by Sodium hydroxide (0.5 M, 5 M). The accepter solution was injected inside lumen of dried membrane through a micro syringe. Specific amount of (24  µl) accepter solution was injected inside lumen of hollow fiber via a BD micro syringe. Specific volume (24  µl) of accepter solution was fixed after several adjustments, for best compatibility with a 7.5 cm hollow fiber, to achieve good repeatability and enrichment. 2.1.2.3 Membrane solvent: Membrane containing accepter solution was dipped for 15 s (Approximately) into the organic solvent (pure DHE or topo% solutions in DHE), to impregnate the fiber with organic solvent and to establish a membrane phase. The solvents, immobilized in the pores of hollow fiber were; pure DHE, 1% topo in DHE (w/v), 5% topo in DHE (w/v), 10% topo in DHE (w/v), 15% topo in DHE (w/v) and 19% topo in DHE (w/v). All solutions (topo in DHE) were prepared and mixed by manual shaking, although 15% topo in DHE and 19% topo in DHE solutions were prepared by vigorous shaking and were put inside sonicator for efficient mixing. 2.2. Sample preparations: All primary solutions were prepared in methanol. Primary solutions were prepared by transferring specific weight of analytes to a sample vial, having air tight caps. This solution was diluted with methanol to prepare a solution of concentration (100 ÃŽ ¼g/ml). Table 2.1 represents properties (physical, chemical) of analytes. A (abbreviation) name was given respective to TMS ester of each analyte, new name consists of three words only. Molecular weight (Mw), Molecular (Molec) formula, Source (chemicals were purchased from), pKa values of individual analytes (dissociates in water) and purity (as labeled on each chemical) of each analyte is listed in Table 2.1. Table. 2.1- Analytes source (purchased from)and purity. Sr. No Chemical name Abbreviation Mw Molec formula Purchased from pka. Values Purity (%) 1 Malonic Acid Mal 104.06 C3H4O4 Aldrich 2.83, 5.69 (36) 99 2 Succinic Acid Suc 118.09 C5H6O4 Fluka 4.19, 5.48 (36) 99.9 3 Glutaric Acid Glu 132.04 C5H8O4 Aldrich 4.34, 5.42 (36) 99 4 Adipic Acid Ad 146.14 C6H10O4 Fluka 4.34,5.44 (36) 99.5 5 Pimelic Acid Pim 160.17 C7H12O4 Aldrich 4.48, 5.42 (36) 98 6 Suberic Acid

Should Gambling Be Legalized Essay -- essays research papers

Should Gambling Be Legalized?   Ã‚  Ã‚  Ã‚  Ã‚  Gambling is the risking of money or other possessions that’s mostly depends on chance and luck, even tough some forms of gambling may be influenced by skills. Some stereotypical forms of gambling are found in casinos they include: roulette, blackjack, craps, poker, slots and many others. Nevertheless, there are other forms of gambling that people would not even consider them so such as playing the lottery. Gambling has been very popular since the beginning of time when people would use instruments of divination in order to try to predict the outcomes of important events such as wars. These devices were very similar to the ones we use for gambling today (Gambling). Gambling continues to be quite popular in today’s society. Nevertheless, many people bel...

How are women portrayed in The Millers Tale Essay

The Miller’s Tale was written and is set in medieval England, a time when women had much fewer rights than men, and were more or less just owned by their fathers, and then by their husbands when they got married. 17th century United States in The Crucible has a slightly different society but also has the similar male dominance. The Handmaid’s Tale is set in a dystopian future where women are also heavily dominated by men, but in a completely different way. This essay is about the ways that women in general are portrayed and perceived in these three stories, as well as touching on the characters of the individual women in these tales. The Miller’s Tale is one of the stories from the Canterbury Tales series, all written in poetic form, by Geoffrey Chaucer. These tales in the series are all told by different pilgrims, who are also fictional, so this uses a story-within-a-story literary device. Their tales are part of a contest to entertain each other on their pilgrimage from Southwark to Canterbury Cathedral. In The Miller’s Tale, it is the miller’s turn to tell a tale, and he tells the story of a devious young student called Nicholas, who is attracted to the much younger wife of a carpenter, his neighbour, and plots a cunning plan to sleep with her. He does this by telling the dim and simple carpenter that a flood is coming, and that he must tie some tubs to the ceiling of his home for the three of them in order to keep them safe. Whilst the carpenter is away at work on these orders, Nicholas takes the carpenter’s wife Alison downstairs and manages to seduce her until she very willingly has sex with him. Alison from The Miller’s Tale is eighteen years old, and described as passionate and highly attractive. Her faithfulness in marriage to her husband is very questionable when she allows herself to be easily taken in by this other man, her neighbour, and commits adultery with him without much care for her own husband. Near the beginning of the Miller’s Tale, there is a clear, physical description of Alison, being a lively woman who might want to have an affair. For she is â€Å"wilde and yonge†, meaning that her behaviour is rather uncontrolled, and her older husband is jealous and possessive of her. The miller describes her as having a â€Å"body gent and smal† as a weasel’s, meaning that she has an attractive slim figure, and that suggests that she is also a very sly character just like a weasel. Alison is also vain and very concerned about her appearance. She is selfish and cares more about herself than of other people, and she does not even think much of all the men that take a strong liking to her. She has established herself as a not at all a likeable character in this story. The fact that she sleeps with Nicholas right in her very own marital home, whilst her own husband is just upstairs at work at the very same time, must show how very daring she is, because he could have easily come downstairs and catch them in the act. But it could also mean that she does not actually really mind or care much about the carpenter’s feelings or whether he knows that she is being unfaithful to him or not. We feel some sympathy for the carpenter, who is being conned like this by two people, as well as being cheated on by his wife and having to bear the shameful title of a â€Å"cuckold†. Alison is certainly one to take risks in exchange for her own selfish sexual desires, going against the female stereotypes of the time by being rebellious and free-spirited and instead of being faithful and modest like a woman should be in her time. Alison from The Miller’s Tale is a lot like Abigail Williams from The Crucible. They are similar ages, and are both selfish and sexually immoral women who both have illicit sexual affairs and go against societal and moral rules that are expected of them for their own personal gain and pleasure. Also, neither Alison nor Abigail show any shred of remorse for their sinful actions. Where Alison goes behind her albeit dim husband’s back to sleep with her neighbour Nicholas, she is in turn fulfilling his desiring lustful plan. Sex outside of marriage was very wrong in her time, let alone committing adultery. Alison might have just married the carpenter for the sake of security, since he is described in the story as a â€Å"rich gnof†, but obviously cannot control her extramarital sexual urges and is very open to acting on them whenever the chance arises. Seventeen year old Abigail betrays her position as a house servant in the Proctor’s home by having an affair with John Proctor whilst he is still married to his kind wife Elizabeth, who happens to be ill at the time the affair occurs. However, there is much more to Abigail than involvements in adultery, as this leads to her seemingly falling in love and becoming obsessed with John Proctor. She says to him in Act One before the trials: â€Å"I know how you clutched my back behind your house and sweated like a stallion whenever I come near†¦ It’s she put me out, you cannot pretend it were you. I saw your face when she put me out, and you loved me then and you do now†. Abigail has seriously formed an impression that John is just as infatuated with her and she is with him, even though he constantly denies it and tells her she is speaking a â€Å"wild thing†. So at the very start of the play, she is casting a spell to kill Elizabeth so that she can be out of the way for herself and John to be together, as she believes that Elizabeth is the only person in her way of having John. We can sympathise a little with Abigail, as we know she has had a very troubled past. She is an orphan, who had watched both her parents being viciously murdered by Indians one night a long time ago. She reveals this in Act One, after ordering the girls to lie about their activities in the woods, she viciously threatens to get them in the night, and in her own words says â€Å"you know I can do it: I saw Indians smash my dear parents’ heads on the pillow next to mine, and I have seen some reddish work done at night, and I can make you wish you had never seen the sun go down! â€Å". This traumatic event that was imposed upon her at such a young age does provide some explanation and understanding as to why her personality seems rather unstable, and why she acts so brutally towards other people. On the other hand, we don’t really know anything about Alison’s past, so we assume she is just a nasty sly character and though her crimes are not as dire as Abigail’s, we do not really have the evidence to feel as much sympathy for her behaviour. Though I think we can like Alison to some extent, as even though we condemn her behaviour, the men in her story are not as admirable as John Proctor so maybe her behaviour does not seem so bad. She even has the advantage of being secure in a marriage, unlike Abigail who is an unmarried orphan living with her uncle. The presentation of Abigail in The Crucible is rather dark and frightening, a good example of this being at the court scene, where she is deliberately causing hysteria by throwing around accusations of witchcraft, and even going as far as pretending to be bewitched by Mary, and getting all the other girls to make believe the very same thing and repeat Abigail’s exact chants and actions.

During The Trimesters Of Pregnancy, Every Change That The

During the trimesters of pregnancy, every change that the baby has, will affect the mother in many different ways. Pregnancy is a nine month process where the mother will experience a lot of changes. On the first trimester, a woman will experience a lot of symptoms as she adjust to the hormonal changes of pregnancy. This process affects nearly very organ on the body. Since the first month the woman body starts to change. The first sign of pregnancy is a missed period. She will not have her period anymore until she give birth. Most pregnancy tests will be positive by the time a woman has missed her period. Also, other early signs of pregnancy include fatigue, which is an extreme tiredness. As well they start feeling bloated and have†¦show more content†¦Women who are prone to acne may experience outbreaks. During the first three months of pregnancy women do not usually gain much weight, they usually get about 2 pounds. Those woman who are overweight or underweight may experience a different rate of weight gain. On these cases they should talk with their health care provider about maintaining a healthy weight throughout the nine months (Parenthood). By the second trimester, some of the unpleasant effects of early pregnancy may lessen or disappear as the body adjust to its changing hormone levels. After the unpleasant nausea of the first one, it appear like a wonderful stage of peace. Sleeping may get easier and energy levels may increase. This trimester of pregnancy often brings feelings of comfort and sense of a renewed well-being. However, there is still a lot of pregnancy symptoms are approaching on the horizon. Some of the early signs and symptoms of pregnancy begin to be relieved during the fourth month. Nausea is usually reduced. But other digestive problems like heartburn and constipation, may be troublesome. Breast changes will continue growing, soreness, and darkening of the areola, also usually continue. It’s very common at this time pf the pregnancy that women are going to have shortness of breath or to breathe faster, which is very uncomfortable. The blood flow increased and may lead to unpleasant pregnancy symptoms, such as bleeding gums, nosebleeds, orShow MoreRelatedPregnancy : The Best And Worst Experience Essay1392 Words   |  6 PagesShowing Trimester One Who is Boss Being a parent is both a blessing and a curse, the best and yet most hardest thing in the world to do. Even when a baby is not planned, and in a lot of cases they are never planned they are still a blessing. Many women have said that even though their little one was not planned, their baby has changed their world in many ways. Some women even said that having that unexpected baby saved their lives. Being pregnant is also the best and worst experience. Some womenRead MoreA Brief Note On The Stages Of Pregnancy1398 Words   |  6 Pages1123-428 29 January 2016 Three Stages of Pregnancy Congratulations! You just found out that you are going to be a new mother. I am sure that you are experiencing a lot of emotions right now, such as excitement and fear. Pregnancy can be extremely overwhelming, regardless of whether it is your first, second or even fifth. â€Å"Each year there are approximately 4 million births in the U.S.† (Elbel par.2). Not only will the new mother-to-be go through these trimesters but the unborn child as well, which weRead MoreThe Experience of Becoming a Mother 1807 Words   |  7 Pageswomb during the pregnancy time can make women enjoy the exciting moment and help in reduce the nervous of giving birth. According to Trish (2004), pregnancy is a special time and a healthy life event. During pregnancy time, women also go through every emotional phase in her life time. A pregnancy woman’s body also will undergoes some physical change. The changes is important to assist for fetal development. There are three stages of pregnancy which a re the first trimester, second trimester and thirdRead MoreConsiderations For The Pregnant Patient1663 Words   |  7 Pagesused occasionally. During her pregnancy she has continued to try to stay active. Rather than going on long runs, she tries to walk every morning. Sarah also enjoys going on walks with Erik and their dog, Duke, after work. On the weekends, they have been remodeling the basement of their home. Sarah is very creative and has been making most of the home decor. She has been staying away from the heavy machinery and letting Erik to the manual improvements. Throughout her pregnancy, Sarah has found thatRead MoreOklahoma s Step Toward Change944 Words   |  4 PagesOklahoma’s Step Towards Change in Abortion Today, teen pregnancy is as high as ever. Shows like â€Å"16 and Pregnant† and â€Å"The Secret Life of an American Teenager† are centered on young girls’ journeys as teen moms, in hopes that viewers will become more aware about safe sex preventing teen pregnancy. Reducing teen pregnancy also reduces the rate of abortions. Teenagers are the ones who are most likely to get an abortion, since they are the ones that are the least prepared to be mothers. RegardlessRead MoreAbortion, The, And Abortion953 Words   |  4 PagesWhether people support or are against abortion, few actually know about the abortion process. Have you ever heard of suction aspiration or prostaglandin chemical abortion? Those are two of the various methods that are performed in the different trimesters of pregnancy. According to writer Steven Ertelt of LifeNews.com, Oklahoma’s abortion laws are restrictive compared to other states, especially with the passage of the Oklahoma Unborn Ch ild Protection from Dismemberment Abortion Act, or HB 1721, in AprilRead MoreTeenage Pregnancy : The Highest Rate Of Teen Pregnancy1666 Words   |  7 PagesTeen Pregnancy The United States has the highest rate of teen pregnancy of most industrialized nations. According to a 2014 article, in 2013 nearly 273,105 babies were born to women aged 15-19 years, for a live birth rate of 26.5 per 1,000 women in this age group. This is a record low for U.S. teens in this age group, and a drop of 10% from 2012. Birth rates fell 13% for women aged 15–17 years, and 8% for women aged 18–19 years. Still, the U.S. teen pregnancy rate is substantially higher thanRead MoreWomen Involved With Intimate Partner Violence1441 Words   |  6 Pagesof IPV in women during pregnancy using the Abuse Assessment Screen in various countries during different stages of pregnancy (Bunn et al., 2009; Hellmuth, Gordon, Stuart, Moore, 2013; Keeling Mason, 2011; Lutgendorf, Thagard, Rockswold, Busch, Magann, 2012; Massumi Okada, Komura Hoga, Vilela Borges, Sartori, Aparecida Belli, 2015; Roelens, Verstraelen, Van Egmond, Temmerman, 2008). In two of the studies, the population studied were pregnant women in the first trimester and postpartum (HellmuthRead MoreFactors Affecting The Rate Of Protein C ( An Anti Coagulation Factor )1099 Words   |  5 Pagesanti-coagulation factor).16 ââ€"‹ Coagulation changes are most pronounced in the third trimester.16 ââ€"  Effect on Dental Treatment o The patient’s hypercoagulable state places them at increased risk for thromboembolic disease.17 Because of this, there is an increased risk that the patient may have a thromboembolism while in the dental office. (See the following section titled â€Å"Thromboembolic Disease† for additional information.) Thromboembolic Disease ââ€"  Background Information ââ€"‹ Changes in clotting factors place allRead MoreSymptoms And Symptoms Of Nausea1221 Words   |  5 PagesWhat Does Nausea Feel Like One of the early signs of pregnancy is nausea, and many expectant mothers experience nausea during early pregnancy. Nausea is usually a common symptom throughout the first trimester or even longer, and it is typically the most complained about symptom that women report. Fortunately, even though nausea can be terribly uncomfortable, it is not harmful to you or your baby. Not surprisingly, first time mothers want to know, what does nausea feel like? What Does Nausea Feel