Effects of Atmospheric Aerosols on Human Health

Effects of Atmospheric Aerosols on Human HealthAbstractA naughtyly Sensitive (LOD 0.04-0.4 ng/ml) method is unquestionable for detection and quantification of panellingic complexs (C3 -C10) containing mono and dicarboxylic acidulouss on GC-MS. These compounds (C3 -C10) existed in trace amount, as level-ranking constitutive(a) aerosols i.e. burning(preno minal) constituents of Aerosols. tissue layer declivity technique was utilised for selective enrichment (1-4300 times) of bottom compounds. Good repeatability (RSD% 10%) from selective complete material body (10% TOPO in DHE) was achieved with troika phase HF-LPME. Aerosols containing patterns, after Ultrasonic Assisted extraction were spy 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 customd for separation, detection and quantification of target compounds.Method r ipening and Application -hollow fiber Supported facile membrane extraction of Fatty acids (C3-C10) containing mono and dicarboxylic acids and Detection of aerosols Samples after ultrasonic back up extraction.1. Introduction blow of Atmospheric aerosols on human health and effect on radioactive stableness in Earths atmosphere is getting grandeur promptly a days and this phenomenon has been well chthonicstood. 1. Atmospheric aerosols roll in the hay harm respiratory and cardiovascular system of human.Impact of Secondary organic aerosols as biogenic and anthropogenetic antecedent is identified (Adams and sinfold, 2002) 1, 17. Low molecular dicarboxylic acids (C3-C9) atomic number 18 as well vital tracers of SOA 2. Short twine roly-poly acids argon prime as secondary organic aerosols which atomic number 18 also supposed to derive from long chain butterball acids 1. Importance of organic aerosol has been well established now a days and carboxylic acids are of great int erest 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 award in aerosol contains to a greater extent than 90% of tropospheres aerosols 5, 15.Dicarboxylic acids found in nature as polymeric compounds such as suberin and attenuatedin 3. Short chain dicarboxylic acids are found in vegetables Siddiqui, 1989 and in soil containing small 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 cyclo poisonousity and antineop operateic activities 18.Many analytical techniques are employ to determine the base of SOA so keeping in view these techniques naked as a jaybird method f or determination of fatty acids (common in SOA) has been developed. Membrane extraction is utilise in this method collectable to its increasing importance for richly 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 differential coefficients can be utilize to make polymers and their condensation with diols in source 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 DescriptionProperties (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 stray. These target analytes are extracted through Liquid phase micro extraction and discover 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 AcidAdipic acid is a product of lipid per oxidation. Adipic acid does not undergo hydrolysis in the environment perhaps receivable to the lack of hydrolysable functional groups (Harris 1990) 5.1.1.2- Malonic AcidMalonic 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 otherwise 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 beetroot rots as a Calcium salt 42.1.1.3- Succinic AcidSuccinic acid is found in atmosphere as water supply 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 acidGlutaric acid is found as SOA in aerosols 8. Glutaric acid is sparingly soluble in water 41, can be apply to prepare a plasticizer for polyester 41.1.1.5- Pimelic AcidPimelic acid is a last dicarboxylic acid relative to carbon number which has IUPAC name. Derivatives of Pimelic acid are use 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 AcidSuberic 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 AcidAzelaic acid is an important constituent of secondary organic aerosols because it produces short chain fatty acids upon icon 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 AcidCis-pinonic acid is also produced in atmosphere by photo oxidation of -pinene in the existence o f Ozone 30.1.1.9- Pinic AcidPinic acid is derivative of -pinene. Pinic acid can be generated by photo oxidation of -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 Acid4-Hydroxy benzoic acid is exists as crystals. It is used to derive parabens and can be used as antioxidant 41.1.1.11-Phthalic AcidPhthalic acid is an aromatic dicarboxylic acid it is found as white crystalline state in pure(a) 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 Auto officious 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 sample s. A theoretical description is given in section 1.2 for 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 ExtractionUltrasonic 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, dis declaration and sample purification. When Ultrasonic technique is used for financial aid in extraction, this assistance in extraction is called Ultrasonic assisted extraction (UAE) 23.There are many advantages by using UAE because it require less organic solvents ,non destructive, less dearly-won and less time consuming comparative to other sample preparation techniques like soxh let 21.The normal range of ultrasound frequencies used in laboratory ranges from 20 KHz to 40 KHz. Use of UAE is simple. A sample resolving power in spite of appearance a vessel in an appropriate solvent can be place inner(a) ultrasonic bath at desired temperature and sound waves stir the sample 20.The mechanism of US is as 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, opus in solid sample both longitudinal and transverse waves can be produced 19. When UAE is utilized it sum ups speed of mass transport by vibration of mechanical transport from the sample matrix through a exhibit called cavitation 21.1.2.2- Theory of Ultrasonic Assisted extractionThere are 2 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 hardheaded use in analytical chemistry.1.2.2.1- Physical aspects of UAEDuring Ultrasonic assisted extraction, a talk 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 extravasate is difficult 21.Ultrasonic metier produces cavitations in a liquid sample during extraction (UAE). Two types of US cavitation is produced known as short-lived cavitation (produce transient bubble) and 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 UAEWhe n 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 unhorse 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 (normally 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 fatuous fiber membrane extractionTwo 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 TechniqueWhen 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 triplet 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 desiccate GC vial along with derivatizing reagent. After derivatization these samples were injected into a Gas chromatographic system.1.3.2 Basic Principle of LPMEBasic principle is same for all LPME techniques ( devil 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 foretoken (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 regulars. 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= D .Korg/sample / a. Korg/accepter.1.2In 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 ratios among Organic phase and sample phase and between accepter phase and organic phase respectivel y 22. D and a are the removable split 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 self-reliant from partition ratio of stationary liquid membrane in three phase lpme i.e. it depends mainly on ionization of analytes in sample 22.Extraction strength (E) can be calculated from Eq. 1.322.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 rati os are directly dependent on partition coefficients, so by increasing the partition ratios efficiency can be meliorate 22. Partition coefficients can be amend 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 deepen in LPMEEnrichment factor (Ee) for three phase LPME is given in Eq. 1.4.Ee = C accepter/C initial= V sample. E / V accepter .. 1.4In 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 () is given in Eq. 1.5 24. = AH/ A-AH = 1/1+10(pH-pKa) .. 1.5In 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 .. 1.6s is equal to 1 for acidic ana lytes (Eq. 1.6). pKa is dissociation constant and pH refers to donor or accepter solution(Eq. 1.6) 24.C = D .Cs a CA.KA/KS 1.7Eq. 1.5-1.7 are derived from Henderson-Hasselbalch relation, in this equation represents the extractable fraction of analytes 24.The driving force for the extraction in neutral conditions of three phase LPME is the concentration gradient (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 extractionThree end points are unremarkably considered for extraction 22.1. thoroughgoing extraction. 2. Kinetic extraction. 3. Equilibrium extraction.1.3.4.1 Exhaustive extractionExhaustive end point is the specific end point (time), when all amount of analytes are exhausted (which can be practical ly possible) present in donor 22. In this practical diploma work, Exhaustive end point will be utilise 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 economic value 12. Mass switch between organic phase and liquid phase is dependent on concentration gradient 12. Enrichment factor can be improved by increasing the value of D preferably close to unity and decreasing the value of A to zero. Such conditions for the D and A determine are called 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 Difference set the value of A to 0.0005, at this point accepter can capture all analytes. At this set value (A), enrichment factor increases line arly with time 12. Peak time of enrichment factor, when other para metres 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 slump again 12.1.3.5 Rate of LPMETwo 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 (C) is approaches to zero described in Eq. 1.8 13, 24.Ee (max) = (C a / C d) max= D/A .. 1.8In 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 K.D m /h m .. 1.9In Eq. 1.9 K is partition coefficient, Dm is membrane diffusion coefficient and h m is the thickness of membran e 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 lpme24Concentration 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 determined back to donor. So this trapping of analytes due to pH adaption is called direct trapping. For high enrichment purpose, pH of accepter is usually adjusted 3.3 pH units higher than the pKa values of acid ic 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 phaseChoice 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 org anic 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 sampleExtraction 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 group is shown in Fig. 1.13.1.3.10Volume of donor and acceptor solutions.Volume of donor and accepter solution is very important because predisposition can be improved by proper volume alteration of accepter solution. Volume of accepter so lution 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 tolerance 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 Analytes1.4.1-GC-MS analysisGC-MS is a powerful detection technique for environmental trace analysis due to its high sensit ivity 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. run down 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. DerivatizationTwo 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 system14. Both derivatizing reagents are applied separately and compared prior to GC-MS analysis.1.5.1- SilylationAnalytes 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 gro ups 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. Method2.1 Membrane extractionThree 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 ExtractionHollow fiber Accurel PP polypropylene (Q3/2) is purchased from Membrana (Wuppertal, Germa ny). The wall thickness of membrane is 200 m, Inner diameter 600 m and pore size is 0.2 m. 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 l (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 Extraction2.1.2.1 Donor solutionThe pH donor solution was adjusted to 2. whole 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 solutionAccepter 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 l) accepter solution was injected inside lumen of hollow fiber via a BD micro syringe. Specific volume (24 l) of accepter solution wa s 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 solventMembrane 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 preparationsAll primary solutions were prepared in methanol. Primary solutions were prepared by transferring specific weight of analytes to a sample vial, having air nigh caps. This solution was diluted with methano l to prepare a solution of concentration (100 g/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. NoChemical nameAbbreviationMwMolec formulaPurchasedfrompka. ValuesPurity(%)1Malonic AcidMal104.06C3H4O4Aldrich2.83, 5.69 (36)992Succinic AcidSuc118.09C5H6O4Fluka4.19, 5.48 (36)99.93Glutaric AcidGlu132.04C5H8O4Aldrich4.34, 5.42 (36)994Adipic AcidAd146.14C6H10O4Fluka4.34,5.44 (36)99.55Pimelic AcidPim160.17C7H12O4Aldrich4.48, 5.42 (36)986Suberic Acid