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HCT 204: CHEMICAL INSTRUMENTATION 1
EXPERIMENT 1:
Determination of the distribution of trace heavy metals in urban roadway dust in Gweru.
INTRODUCTION
Dust is useful as an indicator for pollution. Various parameters can be measured in dust samples chief among which are the heavy metals like Pb, As, Cu, Zn, Ni, Fe, Mn, Al to name a few.When introduced into the human body by inhalation, ingestion or through diffusion via the skin(in the form of salts like ZnCrO4 ) these metals cause various maladies ranging from skin lesions through gastro tract problems to severe brain damage.
People working in dusty environments are exposed to high levels of various toxic metals. The dust that enters the body through inhalation is mostly that which is airborne. In protected areas like open verandas and shades at garages, flea markets and bridges , this air borne dust gradually settles down at a rate dependent on their densities. For dust particles of similar composition, the density should vary directly with particle size. Small particles are air –borne for longer periods than larger ones and end up being inhaled into the human body.
The ease with which dust samples can be taken enhances its use as a pollution index. It is found almost everywhere and can be sampled using very simple equipment.
Dusts along roadways are due to a number of sources. Because of the presence of metals in the earth’s crust, the existence of these metals in roadway dust is to beexpected.Automobiles contribute direct exhaust emission and tyre wear particles which probably come from body rusting and ablation from the interior of the exhaust system. It is apparent that streets with high traffic densities should also have high metal levels.
The construction and demolition of structures or buildings that has been painted with heavy metal –based paints contributes a wide range of metal s into the vicinity of roadways. Some industrial activities like welding, spray painting, metal casting emit toxic metals into the environment.
LITERATURE REVIEW
A literature review is a general survey of the work that has been carried out elsewhere and published in journals by other workers on levels of trace heavy metals in roadway dust.
You are supposed to carry out a literature review and report your findings in less than two pages. Remember it is very important to take note of references in your literature review.
REAGENTS AND CHEM ICALS REQUIRED
Analytical grade (AR) reagents are to be used throughout.
Nitric Acid SG 1.40
Perchloric Acid 72% (w/w)
Hydrochloric Acid SG 1.34
Standard solutions for various metals (1000ppm, spectrosol)
PROCEDURES
Sampling
Identify a sampling site of your choice and prepare an appropriate sampling plan.
Carry out your sampling according to your sampling plan by scooping the dust with clean spatulas into well labeled polythene bags and tie with strings.
Samples should be collected at 2m intervals starting from the edge of the road going away from the road.The total number of samples to be collected should be enough to allow one to draw a good graph of the metal distribution.
Standard Solution Preparation
Using 1000ppm commercial stock solutions transfer 10ml into a 100ml volumetric flask and dilute to the mark with distilled water. This gives a 100ppm working standard for each metal.
Prepare from the working standard appropriate calibration standards by serial dilution measuring 1ppm; 2ppm; 4ppm; 6ppm; and 8ppm. Prepare composite standards. As and Hg standards should be prepared according to the requirements of the methods used to analyse these metals i.e. hydride vapour generation and mercury vapour generation.
Sample Treatment
- Air-dry the dust samples at room temperature, i.e 24-30oC.
- Weigh the dry sample on an analytical balance and sieve the samples through a series of stainless steel of diameter 600,400,200,100 and 50 microns.
- Weigh each fraction and put it in a well labeled bag.
- To 0.4000g portions of the prepared dust in 250ml beakers add distilled water to form a sherry.
- Add HNO3 (5ml) and HCLO4 (0.5ML)
- Heats the mixture on a hot plate until a residue remains. Add 2M HCL (2.5) to the residue and transfer quantitatively the contents to a 100ml volumetric flask bringing to the mark using distilled water.
- Analyse the samples by Flame AA or Graphite furnace AA depending on the level of each element in the samples. Use the following instrumental condition in your analysis by FAA.
METAL / WAVELENGTH(nm) / BAND PASS(nm) / LAMP CURRENT(mA) / FLAME TYPE
Cd / 228.8 / 0.5 / 4 / Oxidising
Cr / 357.9 / 0.2 / 5 / Reducing
Cu / 324.7 / 0.5 / 4 / Oxidising
Fe / 248.3 / 0.2 / 5 / Oxidising
Mn / 279.5 / 0.2 / 5 / Oxidising
Ni / 232.0 / 0.2 / 4 / Oxidising
Pb / 217.0 / 1.0 / 5 / Oxidising
Zn / 213.9 / 1.0 / 5 / Oxidising
Express each fraction as a percentage of the bulk sample and the metal levels as mg metal/kg dry sample.
Draw graphs showing the relationship between;
- Weight of fraction and particle size
- Metal levels and particle size
- Metal level and distance from the road
Discuss and conclude your results.
References
References are expected to come from your literature review.
EXPERIMENT 2:
Ultraviolet spectrophotometric determination of Aspirin, Phenacetin and caffeine and Anadin tablets using solvent extraction.
PRINCIPLE
Caffenol and Anadin tablets are a mixture of aspirin, phenacetin and caffeine. Each of these substances has characteristic absorption in the ultraviolet region, with the principle maximum lying at 227 for aspirin, 275nm for caffeine and 250nm for phenacetin.In the procedure, a powdered tablet is dissolved in methylene chloride and the aspirin is separated from the phenacetin and caffeine by extracting it into aqueous sodium bicarbonate solution. The separated aspirin is back –extracted into methylene chloride by acidifying the aqueous layer and is then measured spectophotometically at 277nm. The phenacetin and caffeine that remain in the original methylene chloride layer are determined in a mixture as described in analytical texts in Chapters on UV/VIS spectrometry according to Beer’s law. According to Beer’s law, when two absorbing species in solution have overlapping peaks, the total absorbance. A is the sum of two absorbances. For two absorbing species,
A=axbcx + aybcy
OR A= Exbcx + Eybcy
Where A is the total absorbance observed ax and ay are the absorptivities of X and Y, Ex and Ey are the molar absorptivities of X and Y in grams per litre of the solution and b is the path length. For two unknowns, two measurements have to be made and absorbencies at each peak will be,
A1=Ax1 +Ay1 =Ex1bcx + Ey1bcy
And Ax2 + Ay2 = Ex2bcx + Ey2bcy
Concentration of x and y can be obtained by solving these equations.
Equations
AHCO-3 A CH2CI2A (277nm)
PCH2CH2 P (250nm) + C (275nm)
c
Reagents and Chemicals
Provided CH2CI2, 4% (wt /vol) NaHCO3 solution (chilled), concentrated. HCl, 1M H2SO4.
To prepare
Standard solutions. Prepare individual standard solutions of about 100mg/L, 20mg/L and 10mg/L each of aspirin, phenacetin and caffeine in methylene chloride as follows:
Weigh about 25mg (to the nearest 0.1mg) of each, transfer to 250ml volumetric flasks, and dissolve to volume with methylene chloride. Dilute 10 and 5ml of this solution to 50ml in 50ml volumetric flasks to prepare the 20 and 10mg/h solutions, respectively.
PROCEDURE
Weigh accurately and record the weight of one tablet. This should be equivalent to about 220mg aspirin, 160mg phenacetin, and 30mg caffeine. To minimize required dilutions and save on solvents, cut the tablet intoquarters and weigh out one –quarter portion to be analysed. Crush to a fine powder in a beaker. Add, with stirring, 20ml methylenechloride; then transfer the mixture quantitatively to a 60ml separatory funnel, rinsing all particles in with a little more methylene chloride. Extract the Aspirin from the methylene chloride solution with two 10ml portions of chilled 4% sodium bicarbonate to which has been added two drops hydrochloric acid, and then with one 5ml portion water. Wash the combined aqueous extracts with three 10ml portions of methylene chloride (to remove traces of water) into a 50ml volumetric flask and dilute to the mark with methylene chloride. Then dilute further a 1ml aliquot of this solution to 50ml with methylene chloride in a volumetric.
Acidify the bicarbonate solution (aqueous extract), still in the separatory funnel, with 6ml of1MH2SO4.This step should be performed without delay, to avoid hydrolysis of aspirin. The acid must be added slowly in small portions. Mix well only after the most of carbon dioxide evolution has ceased. The pH at this point should be 1 to 2 (pH test paper). Extract the acidified solution with eight separate 10ml portions of methylene chloride and filter through an ethylene chloride wet paper into a 100ml volumetric flask. Dilute to volume. Then, Dilute further a 5ml portion of this solution to 25ml with methylene chloride in a volumetric flask.
Record absorbance versus wavelength curves for the standard solutions and unknown solutions between 200 and 300nm. (This step may be deleted if you do not have a recording ultraviolet spectrophotometer). Does the wavelength of 277nm appear to be the most suitable wavelength for the determination of aspirin? Do the wavelengths of 250 and 275nm appear to be the best wavelengths for the measurement of the absorbance for the mixture of phenacetin and caffeine? Explain. Using the absorbencies of the standard and the unknown aspirin solution at 277nm) calculate the percent aspirin in the caffeine and /or Anadin tablets and the number of milligrams of aspirin per tablet keeping in mind the dilutions.
To calculate the concentrations of phenacetin and caffeine, the absorbencies of phenacetin and caffeine standards and of the methylene chloride extract of the sample must all be read at both 250 and 275nm. /using these absorbencies, calculate the percentage phenacetin and caffeine in the APC tablets and the milligrams of each per tablet. See Chapter 13 for the spectrophotometric determination of mixtures. (G.D Christian, Analytical Chemistry)
Note: Aspirin tends to decompose in solution, and analysis should be performed as soon possible after preparing solutions.
EXPERIMENT 3:
Qualitative Gas Chromatographic analysis of a multicomponent mixture of pesticides.
Introduction and Overview
Gas Liquid chromatography (GLC) is a type of chromatography in which the mobile phase is a gas, such as nitrogen, helium, etc, and the stationary phase in an inert liquid. The sample is usually in liquid form, but is flash vapourised as it is injected into the instrument and is maintained in the gaseous state through the instrument. The major components of a gas chromatograph are shown in block diagram below:
Carrier
Gas
Carrier Gas or Mobile Phase
The carrier gas chosen depends; on the detector to be employed; nitrogen or argon is used with the most popular detector, the flame ionization detector (FID). The carrier gas is supplied at a reduced pressure from a large gas cylinder equipped with a pressure regulator. Often the carrier gas is filtered through tubes containing a drying agent, a molecular sieve and oxygenscrubber to remove moisture, impurities and oxygen from the prior to its entering the column. The flow of controller is needle valve or other device used to control the gas flow rate. In some instruments a rotometer is used to measure the the actual flow rate. A flow rate of 75cm3 per min, is most often used for 6,4mm o.d. columns, while a flow rate of 25cm2 per min is used for 3,2 o.d. Columns.
The injection Part
The sample is introduced into the GC through the injection Part, a small hearted chamber capped with a septum. The sample is introduced by means of a small calibrated syringe. The septum is pierced by the Syringe needle and reseals when the syringe needle is withdrawn /typical volumes injected into packed columns are 1-3ul.
The injection part temperature part temperature should be high enough (e.g. 2200C) to vapourise the sample instantly (i.e. flash vapourisation). Sample vapourisaton should be rapid so that the vaporized sample is swept by the carrier gas into the column as a discrete’’plug’’
The column
The column where the actual chromatographic separation occurs is enclosed in an oven that maintains the desired temperature.
A conventional ‘’packed’’ column is filled with a granular solid support coated with a thin layer of liquid stationary phase. Separation occurs by differences in the distribution of the various sample components between the carrier gas and the liquid stationary phase. (s/p).
The s/p is normal coated evenly on the surface of the solid support with a solution of the liquid phase and then evaporating off the solvent. The solid support must have a uniform pore diameter and a large surface area. These properties are needed to support an adequate coating of stationary liquid phase and provide good contact with the mobile phase. The particles should be of regular shape with good mechanical strength to permit an efficient, well-packed column. The solid support can be made from silica and other materials. Columns can be metal or glass. Glass is preferred because it is inert towards most organic compounds. After packing a column is conditioned before its use by passing carrier gas through it at elevated temperatures to remove volatile impurities.
The Detector
The function of the detector is to sense when a compound is leaving the column and to provide a signal that is proportional to the concentration of the compound of the compound in the carrier gas stream. Several types of detectors areavailable. The FID is the most common and responds to all organic compounds. The detector is heated to the temperature needed to keep the sample compounds from condensing.
The Electrometer
The output from the detector is a very small electric current. This is fed into an electrometer which amplifies and converts the detector output to a voltage that is large enough to be recorded.
The Recorder
The recorder records the voltage from the electrometer as a function of time to give a chromatogram showing the separated sample components as peaks in the chromatogram.
Theory of Gas liquid chromatography
Chromatographic Efficiency
The width of chromatographic peaks is a measure of the efficiency of chromatographic system. The system includes the entire instruments and not just the column. The ability to obtain sharp, narrow peaks is often expressed in terms of a plate number. A large value for a plate number indicates high chromatographic efficiency and excellent separation ability.
The plate number, N, is related to the length of the chromatographic column, L, by the equation,
H = L/N
Where H is the plate height.
Effect of flow rate on chromatographic efficiency:
The Van Deemter equation is the classical statement of the effect of flow rate on chromatographic efficiency. The simplified form of this equation is
H = A + B/u + Cu
Where u is the average linear gas flow rate in cm/sec, and A, B and C are constants. A plot of H versus u shows that H has a minimum value at a certain value of u. This is the optimum value of u for chromatographic separation.
H
------
u opt
Temperature and temperature programming
Temperature is a major factor in adjusting conditions for a satisfactory separation. When sample components elute rapidly and are incompletely resolved, lowering the column temperature will slow the elution and probably improve the peak resolution. If a mixture containing both high and low a component is to be separated, the temperature needed to separate the boiling compounds may slow the separation of the high –boiling components too much. Late eluting peaks will be broader and resolution often poor. In such cases temperature programming can be used. In this technique, the column temperature is increased linearly with time at a preset rate. The more volatile sample components are separated at the lower temperatures while the higher- boiling compounds gradually move at a faster rate through the column so that their peaks appear earlier on the chromatogram.
Principle of method
An acetone extract of the specimen is partitioned between hexane and saturated brine. The hexane, containing the non polar pesticides (organophosphorus, organochlorine and carbaryl), plus fat, is cleaned up on a wood’s column and then examined by the GLC,TLC Brine solution containing the polar pesticides is extracted by ethyl acetate and extract examined by GLC,TLC, e.t.c. If only one type (polar or non-polar) is being sought the full procedure need not be followed, the appropriate parts are selected.
Procedure
Put 10g of soil sample and 10g anhydrous sodium sulphate and 50ml acetone in a250ml beaker and homogenize by stirring. Filter the contents through a fluted filter paper and place on water bath with fan to evaporate to approximately 5- 10ml.
Transfer the residue to a 250ml separating funnel containing 100ml of saturated sodium chloride solution and rinse the beaker with 2x 5 ml hexane and 1 x 5 ml acetone adding the rinsing to the separating funnel. Shake vigorously for 2 minutes, allow the layers to separate and run the aqueous layer into the original beaker. Transfer the hexane to a 250ml separatory funnel.
Re-extract the aqueous layer twice or more, each with 10ml hexane. Wash the combined hexane with 2 x 10 ml brine solution by gentle shaking, and add the washings to the main brine solution, which is then reserved for the extraction of the polar pesticides.
If necessary, dry the combined hexane with a little solid anhydrous sodium sulphate and then decant onto 1.5g celite in a 50ml beaker, washing the sodium sulphate with a little hexane. Evaporate the hexane at low temperature, with occasional stirring until a homogenous hexane-free friable powder is obtained and transfer to a small wood’s column for clean up, Fill in the large wood’s columns with Florasil and saturate with hexane. Elute from the small columns into the large columns using dimethyl sulphaxide (DMSO) (6ml).Elute from the large columns with 50ml hexane containing 15% of ether. Concentrate the eluate to 10ml and reserve for GLC analysis.
Extract the combined brine solution with 3 x 50 ml ethyl acetate (vigorous shaking for 5 minutes) drying each extract successively with the small quantity of anhydrous sodium sulphate and evaporating successively in a 150ml beaker, finally to almost dryness. Dissolve in acetone and make up to 10ml, and reserve for GLC.This solution contains the polar organophosphorus pesticides.
GLC ANALYSIS
It is advisable to inject first on FID, then NPD and possibly after appropriate dilution, on ECD.
Confirmation of identity can sometimes be partly accomplished by injection on more than one column, but if possible, TLC confirmation should be done.
FID – Flame Ionisaton Detector, non selective, detects both polar and non polar pesticides.
NPD- Nitrogen Phosphorus Detector; selective, detects polar pesticides.
ECD- Electron capture Detector; selective, detects non polar pesticides