International Programme on Chemical Safety

Basic Analytical Toxicology


Monographs - analytical and toxicological data (6.12 - 6.23)

6.12 Bismuth

Bismuth (Bi) has industrial uses in pigments and the production of alloys. In medicine, the main applications of bismuth salts, such as bismuth subsalicylate, are for the treatment of gastrointestinal problems like gastritis, peptic ulcer and diarrhoea. As with antimony, arsenic and mercury, bismuth can be detected using the Reinsch test.

Qualitative test

Applicable to urine, stomach contents and scene residues. Reinsch test - see antimony monograph (section 6.5)

Results

Staining on the copper can be interpreted as follows:

purple black - antimony

dull black - arsenic

shiny black - bismuth

silvery - mercury

Selenium and tellurium may also give dark deposits, while high concentrations of sulfur may give a speckled appearance to the copper.

An estimation of the concentration of bismuth in the sample can be made by comparison of the deposit on the copper with that obtained from a solution containing a known concentration of the element.

Sensitivity

Bismuth, about 2 mg/l.

Confirmatory test

Applicable to blackened copper from the test above.

Reagents

1. Aqueous potassium cyanide solution (100 g/l). Take care when using concentrated cyanide solutions.

2. Aqueous sodium sulfite solution (50 g/l, freshly prepared).

3. Aqueous nitric acid (3 mol/l).

4. Quinine/potassium iodide reagent. Dissolve 1 g of quinine sulfate in 100 ml of purified water containing 0.5 ml of concentrated nitric acid (relative density 1.42). Add 2 g of potassium iodide when the quinine has completely dissolved.

Method

1. Place the copper in potassium cyanide solution and allow to stand for 10 minutes.

2. Wash any undissolved stain with purified water and add 1 ml of sodium sulfite solution and 1 ml of dilute nitric acid.

3. Agitate frequently for 5 minutes and add 1 ml of purified water followed by 1 ml of quinine/potassium iodide reagent.

Results

Stains due to arsenic dissolve in potassium cyanide solution, while stains due to antimony and bismuth do not. However, bismuth slowly forms an orange/brown suspension with quinine/potassium iodide reagent.

Sensitivity

Bismuth, about 2 mg/l.

Clinical interpretation

Acute poisoning with bismuth may cause renal damage, encephalopathy and peripheral neuropathy. Neurotoxicity may also occur after chronic treatment with bismuth salts, but is reversible if medication is stopped.

6.13 Borates

Borates are found in household products as either boric acid (H3BO3) or borax (sodium borate, disodium tetraborate, Na2B4O7), and are used in insecticides, fungicides, wood-preservatives, cleaning agents and water softeners. Weak solutions are used in eye-drops, eye- lotions, mouth washes, depilatory agents and other topical ointments. Small children are especially susceptible to borates and deaths have occurred after topical application of boric acid powder for happy rash. Serious borate poisoning in adults is usually the result of improper use. The fatal dose of boric acid or sodium borate in an adult is 7-35 g.

Qualitative test

Applicable to stomach contents and scene residues.

Reagents

1. Turmeric (the spice) solution (10 g/l) in methanol.

2. Aqueous hydrochloric acid (1 mol/l).

3. Aqueous ammonium hydroxide (4 mol/l).

Method

1. Soak strips (1 × 5 cm) of filter-paper in turmeric solution and allow to dry at room temperature.

2. Add 1 ml of dilute hydrochloric acid to 1 ml of sample and soak a strip of turmeric paper in the solution.

3. Allow the paper to dry and then moisten with ammonium hydroxide solution.

Results

A brownish-red colour is obtained initially which intensifies as the paper dries. A change to green-black upon moistening with ammonium hydroxide indicates the presence of borates. Oxidizing agents (including bromates, chlorates, iodates and nitrites) interfere because they bleach turmeric.

Sensitivity

Borate, 50 mg/l.

Confirmatory test

Applicable to stomach contents and scene residues.

Reagent

Carminic acid (0.5 g/l) in concentrated sulfuric acid (relative density 1.83).

Method

1. Filter, if necessary, 5 ml of stomach contents into a 10-ml glass tube.

2. Add 0.5 ml of filtrate or scene residue to a clean tube and slowly add 0.5 ml of carminic acid solution down the side of the tube so that it forms a layer under the sample.

Results

A blue-violet ring at the junction of the two layers indicates the presence of borate. Strong oxidizing agents (including bromates, chlorates, iodates and nitrites) also give positive results in this test.

Sensitivity

Borate, 100 mg/l.

Quantitative assay

Applicable to plasma:or serum (1 ml).

Reagents

1. Aqueous ammonium sulfate solution (40 g/l).

2. Concentrated sulfuric acid (relative density 1.83).

3. Carminic acid (0.2 g/l) in concentrated sulfuric acid.

Standards

Dissolve 0.210 g of boric acid in 100 ml of purified water (borate ion, 2.00 g/l) and dilute with blank serum to give standard solutions containing borate ion concentrations of 20, 50, 100 and 200 mg/l.

Method

1. Add 5 ml of ammonium sulfate solution to 1 ml of sample or standard, vortex-mix, and heat in a boiling water bath for 15 minutes.

2. Centrifuge for 10 minutes and transfer the supernatant to a 10-ml volumetric flask.

3. Shake the precipitate with 2 ml of water, centrifuge as above and again transfer the supernatant to the flask.

4. Make up to 10.0 ml with purified water and mix for 5 seconds.

5. To 1 ml of the solution from the flask add 5 ml of concentrated sulfuric acid and mix thoroughly.

6. Add 5 ml of carminic acid solution, mix thoroughly and allow to stand for 10 minutes.

7. Read the absorbance at 600 nm against a serum blank (see section 4.5.2).

Results

Plot a graph of absorbance against borate concentration in the calibration solutions and calculate the borate concentration in the sample.

Sensitivity

Borate, 20 mg/l.

Clinical interpretation

Clinical features of borate poisoning include nausea, vomiting, diarrhoea, coma, convulsions and circulatory collapse. Haemodialysis or peritoneal dialysis may be indicated in severe cases. Normally, serum borate concentrations range up to 7 mg/l, but serious toxicity may occur at concentrations of 20-150 mg/l. Death may occur at concentrations ranging from 200 mg/l to 1500 mg/l.

6.14 Bromates

Bromates, such as sodium bromate (NaBrO3), are used as ingredients in hair treatment (home perm) kits and are strong oxidizing agents. The diphenylamine test given below will detect other such compounds, notably chlorates, hypochlorites, iodates, nitrates and nitrites.

Qualitative test

Applicable to stomach contents and scene residues.

Reagent

Diphenylamine (10 g/l) in concentrated sulfuric acid (relative density 1.83).

Method

1. Filter, if necessary, 5 ml of stomach contents into a 10-ml glass tube.

2. Add 0.5 ml of filtrate or scene residue to a clean tube and slowly add 0.5 ml of diphenylamine solution down the side of the tube so that it forms a layer under the sample.

Results

A positive result is indicated by a strong blue colour which develops immediately at the junction of the two layers. A light blue colour will be given by most samples of stomach contents owing to the presence of organic material. Since all strong oxidizing agents are rapidly reduced in biological samples, the test should be performed as soon as possible after receipt of the sample.

Sensitivity

Bromate, 10 mg/l.

Confirmatory test

Applicable to urine, stomach contents and scene residues.

Reagents

1. Aqueous nitric acid (2 mol/l).

2. Aqueous silver nitrate solution (10 g/l).

3. Aqueous sodium nitrite solution (50 g/l, freshly prepared).

4. Concentrated ammonium hydroxide (relative density 0.88).

Method

1. To 1 ml of sample add 0.2 ml of dilute nitric acid and 0.2 ml of silver nitrate solution and mix for 5 seconds.

2. If a precipitate forms (owing to the presence of halides), centrifuge in a bench centrifuge for 1 minute and retain the clear supernatant.

3. Add more silver nitrate solution, drop by drop, to ensure the complete removal of any halide, and then add 0.2 ml of sodium nitrite solution.

4. If a precipitate forms add 0.2 ml of concentrated ammonium hydroxide.

Results

A cream precipitate sparingly soluble in ammonium hydroxide indicates the presence of bromate. Iodate reacts similarly to halides in this test (see step 3).

Sensitivity

Bromate, 10 mg/l.

Clinical interpretation

Acute bromate poisoning may cause nausea, vomiting, diarrhoea, abdominal pain, confusion, coma and convulsions. Methaemoglobinaemia is often produced and this may be indicated by dark chocolate-coloured blood (see section 3.2.2). Blood methaemoglobin can be measured but is unstable, and the use of stored samples is unreliable. Treatment is symptomatic and supportive.

6.15 Bromides

Salts such as sodium bromide (NaBr) are sometimes still used as sedatives and anticonvulsants, and are also employed in photographic processing. Methyl bromide is used as a fumigant in ships' holds and grain silos, and is partly metabolized to bromide ion in vivo. Brominated sedatives such as carbromal also give rise to inorganic bromide when metabolized. The qualitative test given below serves to indicate the presence of inorganic bromide or iodide, and the appropriate confirmatory tests must then be used.

Qualitative test

Applicable to urine, stomach contents and scene residues.

Reagents

1. Aqueous nitric acid (2 mol/l).

2. Aqueous silver nitrate solution (10 g/l).

3. Concentrated ammonium hydroxide (relative density 0.88).

Method

1. Add 0.1 ml of nitric acid to 1 ml of clear test solution, mix for 5 seconds and add 0.1 ml of silver nitrate solution.

2. Centrifuge to isolate any significant precipitate, decant and treat with 0.1 ml of concentrated ammonium hydroxide.

Results

A white precipitate soluble in ammonium hydroxide indicates chloride, an off-white precipitate sparingly soluble in ammonium hydroxide indicates bromide, and a creamy yellow, insoluble precipitate indicates iodide.

This procedure can also be used to test for organobromine sedatives such as carbromal in stomach contents and scene residues. First boil 1 ml of the sample with 1 ml of aqueous sodium hydroxide (5 mol/l) for 5 minutes, cool and neutralize by slowly adding 3 ml of nitric acid (2 mol/l). Then proceed with step 1 above.

Sensitivity

Bromide, 50 mg/l.

Confirmatory test

Applicable to urine, stomach contents and scene residues.

Reagents

1. Saturated fluorescein solution in aqueous acetic acid (600 ml/l).

2. Concentrated sulfuric acid (relative density 1.83).

3. Potassium permanganate (solid)

Method

1. Soak a strip of filter-paper in fluorescein solution.

2. Add about 50 mg of potassium permanganate to 2 ml of test solution in a 10-ml test-tube.

3. Add 0.2 ml of concentrated sulfuric acid and hold the fluorescein-impregnated filter-paper in the mouth of the tube.

Results

Bromide is oxidized to free bromine. This reacts with the yellow dye fluorescein to give eosin (tetrabromofluorescein) which has a pink/red colour.

Sensitivity

Bromide, 50 mg/l.

Quantitative assay

Applicable to plasma or serum (2 ml).

Reagents

1. Aqueous chloroauric acid. Dissolve 0.5 g of chloroauric acid (gold chloride, HAuCl4ÊxH2O) in 100 ml of purified water.

2. Aqueous trichloroacetic acid (200 g/l).

Standards

Dissolve 1.29 g of sodium bromide in 500 ml of purified water (bromide ion 2 g/l). Prepare serial dilutions in purified water containing bromide ion concentrations of 0.2, 0.4, 0.6, 0.8, 1.2 and 1.6 g/l.

Method

1. Add 6 ml of trichloroacetic acid solution to 2 ml of sample in a 10-ml test-tube, vortex-mix for 30 seconds and allow to stand for 15 minutes.

2. Centrifuge in a bench centrifuge for 5 minutes and filter the supernatant through phase-separating filter-paper into a clean tube.

3. Add 1 ml of chloroauric acid solution to 4 ml of the clear supernatant and vortex-mix for 5 seconds.

4. Record the absorbance at 440 nm against a purified water blank (see section 4.5.2).

Results

Construct a calibration graph of bromide concentration against absorbance by analysis of the standard bromide solutions, and calculate the concentration of bromide ion in the sample. The calibration is linear for concentrations from 25 mg/l to 2.5 g/l. This method is not reliable with specimens that may give turbid supernatants, e.g. postmortem samples.

Sensitivity

Bromide, 25 mg/l.

Clinical interpretation

Following acute overdosage, bromides may cause nausea, vomiting and diarrhoea, but absorption is poor and systemic toxicity is more usual after chronic ingestion or exposure. In such cases, fatigue, irritability, anorexia, abdominal pain, skin pigmentation, visual and auditory hallucinations, delirium, tremor, ataxia and coma may occur.

Normal serum bromide concentrations are less than 10 mg/l but, following administration of bromides in therapy, concentrations of up to 80 mg/l may be attained. Toxicity is usually associated with bromide concentrations greater than 500 mg/l. Treatment is normally symptomatic and supportive.

6.16 Cadmium

Cadmium (Cd) forms colourless salts with chemical properties similar to those of zinc compounds. Cadmium oxide and cadmium salts and alloys are used in products such as nickel-cadmium dry batteries, solder, paint and plastic pigments. Acute poisoning due to cadmium is extremely rare, but chronic toxicity has been noted after occupational exposure and in some instances after the diet or the water supply has been contaminated, as with itai-itai (ouch-ouch) disease in Japan.

There is no simple qualitative test for cadmium that can be performed on biological samples or scene residues.

Clinical interpretation

Chronic exposure to cadmium may lead to renal tubular damage and impaired lung function. Osteomalacia has also been observed in cases where the diet is deficient in calcium. Ingestion of cadmium salts causes abdominal pain, vomiting and diarrhoea, with facial oedema, hypotension, metabolic acidosis, depressed respiration, pulmonary oedema, oliguria and death in severe cases. Treatment is symptomatic and supportive, and may include chelation therapy.

6.17 Caffeine

Methyltheobromine, 7-methyltheophylline, 1,3,7-trimethylxanthine; C8H10N4O2; relative molecular mass, 194

Caffeine is an alkaloid present in tea, coffee, cola and other beverages. A cup of coffee or tea may contain up to 100 mg of the drug. Caffeine is an ingredient of many proprietary stimulant preparations and is also used to treat neonatal apnoea. Metabolic reactions include N-demethylation and oxidation to uric acid derivatives. About 85% of an oral dose is excreted unchanged in urine. Caffeine is an important metabolite of theophylline in neonates, and in adults with impaired drug handling.

There is no simple qualitative test for caffeine, but this compound can be detected and identified by thin-layer chromatography of a basic solvent extract of urine, stomach contents or scene residues (see section 5.2.3). However, caffeine responds only to the acidified iodoplatinate visualization reagent, and sensitivity is poor.

Clinical interpretation

Acute overdosage with caffeine may cause palpitations, hypertension, diuresis, central nervous system stimulation, nausea, vomiting, marked hypokalaemia, metabolic acidosis and convulsions. Treatment is generally symptomatic and supportive.

6.18 Camphor

Bornan-2-one; C10H16O; relative molecular mass, 152

Camphor is a rubefacient and is also used in mothballs. It is obtained by distillation from the wood of Cinnamomum camphora or synthetically, and is metabolized by hydroxylation and excreted as glucuronides in urine. Camphor poisoning is usually due to ingestion of camphorated oil. The fatal dose in an adult may be as little as 4 g.

There is no simple qualitative test for this compound. However, camphor has a strong, distinctive smell and detection of this odour on the breath or in urine may aid the diagnosis.

Clinical interpretation

Ingestion of camphor may cause nausea, vomiting, headache, confusion, vertigo, excitement, hallucinations, tremor and dilated pupils. In severe cases, coma, convulsions and hepatorenal failure may ensure. Treatment is symptomatic and supportive.

6.19 Carbamate pesticides

These compounds have the general formula shown below. Substitution of sulfur for oxygen also occurs, but such compounds generally have low insecticidal activity. Some common carbamates are listed in Table 21.

Carbamates are widely used as insecticides, herbicides and fungicides. Carbamate insecticides inhibit acetylcholinesterase and thus evidence of exposure to such compounds can be obtained by measuring cholinesterase activity (see sections 3.1.5 and 6.30). Herbicide and fungicide carbamates, such as the dithiocarbamates, do not inhibit cholinesterase to any significant degree and are relatively nontoxic in humans. The test described here is based on a general reaction of carbamates with furfuraldehyde in the presence of hydrogen chloride.

Qualitative test

Applicable to stomach contents and scene residues.

Reagents

1. Aqueous hydrochloric acid (2 mol/l).

2. Furfuraldehyde solution (100 ml/l) in methanol, freshly prepared.

3. Concentrated hydrochloric acid (relative density 1.18).

Method:

1. Acidify 1 ml of sample with 0.5 ml of dilute hydrochloric acid and extract with 4 ml of chloroform on a rotary mixer for 5 minutes.

2. Centrifuge in a bench centrifuge for 5 minutes, discard the upper, aqueous layer and filter the chloroform extract through phase-separating filter-paper into a clean tube.

3. Evaporate the extract to dryness under a stream of compressed air or nitrogen at 40°C.

4. Dissolve the residue in 0.1 ml of methanol, apply a spot of the solution to filter-paper and allow to dry.

5. Apply 0.1 ml of furfuraldehyde solution to the spot, allow to dry and expose the paper to concentrated hydrochloric acid fumes for 5 minutes in a fume cupboard.

Results

Carbamates give a black spot. Meprobamate and other non-pesticide carbamates interfere in this test.

Sensitivity

Carbamate, 100 mg/l.

Clinical interpretation

Exposure to carbamates may cause anorexia, abdominal pain, nausea, vomiting, diarrhoea, lacrimation, increased salivation, sweating, anxiety, ataxia and acute pulmonary oedema. Antidotal therapy with atropine may be indicated, but pralidoxime should not be used.

6.20 Carbamazepine

5 H-Dibenz [b,f]azepine-5-carboxamide; C15H12N2O; relative molecular mass, 236

Carbamazepine is widely used as an anticonvulsant. Metabolic reactions include epoxidation to give carbamazepine-10,11-epoxide (which is pharmacologically active), diol formation, hydroxylation and conjugation. Less than 10% of a dose is excreted in urine as the parent compound. The estimated minimum lethal dose in an adult is 5 g.

Qualitative test

Applicable to stomach contents and scene residues.

Reagents

1. Aqueous hydrochloric acid (2 mol/l).

2. Sodium hypobromite reagent. Dissolve 0.5 ml of elemental bromine carefully and with cooling in 5 ml of aqueous sodium hydroxide solution (400 g/l). Prepare freshly.

Method

1. Add 1 ml of dilute hydrochloric acid to 5 ml of sample and 5 ml of chloroform, vortex-mix for 1 minute and centrifuge in a bench centrifuge for 5 minutes.

2. After discarding the upper, aqueous layer, add 1 ml of the chloroform extract to 0.2 ml of sodium hypobromite reagent in a clean tube and vortex-mix for 30 seconds.

Results

A blue-violet colour in the chloroform layer indicates the presence of carbamazepine. This compound and its metabolites can also be detected by thin-layer chromatography of an acidic extract of urine (see section 5.2.3).

Sensitivity

Carbamazepine, 250 mg/l.

Clinical interpretation

Carbamazepine poisoning may cause headache, dry mouth, abdominal discomfort, diarrhoea, constipation, ataxia, nystagmus, diplopia, hypotension, coma, convulsions and respiratory depression. Treatment is generally symptomatic and supportive.

6.21 Carbon disulfide

Carbon disulfide (CS2) is used as a synthetic intermediate, a solvent (especially in viscose rayon manufacture), a grain and soil fumigant, an insecticide, a corrosion inhibitor and in degreasing. Some 50-90% of an ingested dose of carbon disulfide is metabolized and excreted in urine as inorganic sulfate, thiourea, 2-mercapto-2- thiazolin-5-one and 2-thiothiazolidine-4-carboxylic acid (TTCA). Carbon disulfide has a particularly pungent smell. The ingestion of 15 ml may prove fatal in an adult.

There is no simple method for the detection of carbon disulfide in biological specimens other than by smell. However, the method given below can be used to assess exposure and relies on the fact that TTCA catalyses the decolorization of a solution of iodine by sodium azide.

Qualitative test

Applicable to urine.

Reagents

1. Iodine-azide reagent. Dissolve 3 g of sodium azide in 25 ml of purified water, add 50 ml of an aqueous solution containing iodine (24.5 g/l) and potassium iodide (50 g/l), and dilute to 100 ml.

2. Aqueous sodium dihydrogen orthophosphate (110 g/l).

Method

1. Add 0.2 ml of sodium dihydrogen orthophosphate solution to 1.0 ml of test urine and to 1.0 ml of blank urine in separate test- tubes.

2. Vortex-mix for 2 seconds, add 20 µl of iodine-azide reagent to each tube and again vortex-mix for 2 seconds.

Results

The yellow-brown colour (of iodine) is decolorized within 30 seconds in the presence of TTCA at room temperature. It is especially important to analyse the blank urine as well as the test urine in this instance, since urine itself often has a yellow-brown colour.

Sensitivity

TTCA, 10 mg/l.

Clinical interpretation

Carbon disulfide is an excellent solvent for fat, and dermal contact can cause reddening, burning, cracking and peeling of the skin. Acute poisoning from either ingestion or inhalation may give rise to irritation of mucous membranes, blurred vision, headache, nausea, vomiting, coma, convulsions and cardiorespiratory arrest.

Following chronic exposure, peripheral neuropathy, fatigue, sleep disturbance, anorexia, weight loss, depression, intellectual impairment, diabetes mellitus and ischaemic heart disease may occur. Treatment is generally symptomatic and supportive.

6.22 Carbon monoxide

Carbon monoxide (CO) is an important constituent of coal gas, but is not present in natural gas. Nowadays, common sources of carbon monoxide are automobile exhaust fumes, improperly maintained or ventilated gas or fuel oil heating systems, and smoke from all types of fires. Carbon monoxide is also produced in vivo from the metabolism of dichloromethane.

Carbon monoxide is highly poisonous and combines with haemoglobin and other haem proteins such as cytochrome oxidase, thereby limiting the oxygen supply to tissue and inhibiting cellular respiration. The affinity of carbon monoxide for haemoglobin is about 200 times that of oxygen. Thus, severe acute or acute-on-chronic poisoning can occur when relatively small quantities of carbon monoxide are present in the inspired air.

The qualitative test described below is relatively insensitive and is useful only in the diagnosis of acute carbon monoxide poisoning. If a positive result is obtained then either the blood carboxyhaemoglobin (HbCO) or the breath carbon monoxide concentration should be measured without delay. The quantitative method for determining blood HbCO described below relies on the fact that both oxygenated haemoglobin and methaemoglobin (oxidized haemoglobin) can be reduced by sodium dithionite while HbCO is largely unaffected.

Qualitative test

Applicable to whole blood treated with heparin, edetic acid or fluoride/oxalate.

Reagent

Aqueous ammonium hydroxide (0.01 mol/l).

Method

Add 0.1 ml of blood to 2 ml of ammonium hydroxide solution and vortex-mix for 5 seconds.

Results

A pink tint in comparison with the colour obtained from a normal blood specimen suggests the presence of carboxyhaemoglobin. Cyanide may give a similar tint, but acute cyanide poisoning is generally much less common than carbon monoxide poisoning.

Sensitivity

HbCO, 20%.

Quantitative assay

Applicable to whole blood treated with heparin, edetic acid or fluoride/oxalate.

Reagents

1. Aqueous ammonium hydroxide (1 ml/l).

2. Sodium dithionite (solid, stored in a desiccator).

3. A supply of pure carbon monoxide or carbon monoxide/nitrogen.

4. A supply of oxygen or compressed air.

Method

1. Add 0.2 ml of blood to 25 ml of ammonium hydroxide solution and mix.

2. Take three approximately equal portions: x, y and z. Keep portion x in a stoppered tube while the following procedures are performed:

(a) Saturate portion y with carbon monoxide (to give 100% HbCO) by bubbling the gas through the solution for 5-10 minutes. Take care to minimize frothing.

(b) Saturate portion z with oxygen by bubbling pure oxygen or compressed air through the solution for at least 10 minutes to remove all bound carbon monoxide (to give 0% HbCO). Again, take care to minimize frothing.

3. Add a small amount (about 20 mg) of sodium dithionite to each test solution (x, y and z) and also to 10 ml of ammonium hydroxide solution and mix well.

4. Measure the absorbances of solutions x, y and z against the dithionite-treated ammonium hydroxide solution at 540 nm and 579 nm.

Results

The percentage carboxyhaemoglobin saturation (% HbCO can be calculated from the equation:

(A540/A579solution x) - (A540/A579solution z) %HbCO = × 100

(A540/A579solution y) - (A540/A579solution z)

Approximate normal values are:

(A540/A579 solution y) = 1.5, corresponding to 100% HbCO

(A540/A579 solution z) = 1.1, corresponding to 0% HbCO.

Note that the haemoglobin content of blood varies from person to person, and thus the volume of diluent used may need to be altered. A dilution giving a maximum absorbance of about 1 absorbance unit at 540 nm is ideal.

It is important to use sodium dithionite that has been freshly obtained or stored in a sealed container in a desiccator, since this compound is inactivated by prolonged contact with moist air.

This method is unreliable in the presence of other pigments such as methaemoglobin (indicated by a relatively high absorbance in the region 580-600 nm, see Fig. 11). Lipaemic blood specimens may give turbid suspensions which also give unreliable results.

The measurements are performed at the point of maximum difference of absorbance (540 nm, lambdamax HbCO) and the point of equal absorbance (579 nm, isobestic point). The reading at 579 nm is taken on a very steep slope (Fig. 11), and the wavelength is critical. Spectrophotometers with a relatively broad band-pass (4-5 nm) should not be used, since it will be impossible to perform the measurement with the accuracy required. Even if an instrument with a narrow band- pass is available, it is important to ensure that it is accurately calibrated, although the effect of minor variations can be minimized by using the following procedure:

1. Measure the absorbance of solution z (0% HbCO) against the dithionite-treated ammonium hydroxide solution at 540 nm. If the ratio (A540/A579) for 0% HbCO is assumed to be 1.1, the absorbance of this solution at 579 nm can be calculated.

2. Adjust the wavelength setting of the instrument to give this reading if not already attained at 579 nm. Alternatively, the spectra from the three solutions can be recorded using a scanning spectrophotometer, if available, and the measurements performed directly. Examples of the spectra that should be obtained are given in Figure 11. The presence of the twin absorption peaks ("rabbit's ears") is a useful qualitative feature.

Sensitivity

HbCO, approximately 10%.

Clinical interpretation

Features of acute carbon monoxide poisoning include headache, nausea, vomiting, haematemesis, hyperventilation, cardiac arrhythmias, pulmonary oedema, coma and acute renal failure. Cyanosis is commonly absent, so that skin and mucosae remain pink even in the presence of severe tissue hypoxia. Death often ensues from respiratory failure. Late neuropsychiatric sequelae are an increasingly recognized complication.

Treatment consists of removal from the contaminated atmosphere and administration of 100% oxygen via a well-fitting face-mask. Hyperbaric oxygen may be indicated in certain cases, and is especially effective in preventing the development of late sequelae, but facilities where this can be given are rare.

Once the patient is removed from the contaminated atmosphere, carboxyhaemoglobin is dissociated rapidly, especially if oxygen is administered in treatment. HbCO measurements are therefore often unhelpful as an indication of the severity of poisoning except in forensic toxicology. A simple guide to the interpretation of blood HbCO results is given in Table 22.

6.23 Carbon tetrachloride

Tetrachloromethane; CCl4; relative molecular mass, 154

Carbon tetrachloride was widely used as a dry-cleaning and degreasing agent and in fire extinguishers. However, as with chloroform, exposure to carbon tetrachloride frequently gives rise to hepatorenal damage and nowadays usage is largely restricted to fumigation of grain and industrial applications.

Massive exposure to carbon tetrachloride may be detectable in urine using the Fujiwara test, possibly because chloroform is a minor metabolite or contaminant; carbon tetrachloride itself does not react in this test.

Qualitative test

Applicable to urine. Fujiwara test. This test must be performed in a fume cupboard.

Reagents

1. Aqueous sodium hydroxide solution (5 mol/l, i.e., 200 g/l).

2. Aqueous trichloroacetic acid (10 mg/l).

Method

1. To separate 10-ml tubes add 1-ml portions of:

(a) test urine;

(b) purified water; and

(c) trichloroacetic acid solution.

2. Add 1 ml of sodium hydroxide solution and 1 ml of pyridine to each tube, mix gently and fit with a loose stopper.

3. Heat in a boiling water-bath for 2 minutes.

Results

An intense red/purple colour in the upper, pyridine layer indicates the presence of trichloro compounds. The blank analysis excludes contamination with compounds such as chloroform from the laboratory atmosphere. Compounds such as chloral hydrate, dichloralphenazone and trichloroethylene, which are extensively metabolized to trichloroacetic acid, give strong positive results in this test.

Sensitivity

Trichloroacetate, 1 mg/l.

Clinical interpretation

Acute poisoning with carbon tetrachloride is rare. Clinical features include ataxia, nausea, vomiting, coma, convulsions, respiratory depression and cardiac arrhythmias. Hepatic and renal damage commonly occur. Treatment is symptomatic and supportive, although acetylcysteine may protect against liver and kidney damage.