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SALICYLATE INTERFERENCE WITH PLASMA-PARACETAMOL METHOD
1362 tion (maintenance) of the blood-pressure at that time. Nearly everyone has accepted this fact for years-that the sympathetic nervous system, the renin/angiotensin/aldosterone axis, volume, cardiac output, peripheral resistance, and so on are all involved in blood-pressure regulation. However, the demonstration that reduction in the activity of one of these components or regulators lowers blood-pressure does not establish it as a cause of the hypertension. This is true even if control of the high blood-pressure is associated chronologically with the return of the regulator to normal. Such an association was
by Gill et aB.4 and McAllister et al/ High plasmarenin-activity appeared to respond to control of accelerated hypertension by agents which have very little or no direct effect on renin release per se. Thus, the high blood-pressure could very well be the cause, and the raised renin activity the result, rather than vice versa as suggested by Dr Moore and demonstrated
Dr Goodwin. Division of Clinical
CALCIUM FLUX AND CYSTIC FIBROSIS
SiR,—That ionic disturbances are an important factor in cystic fibrosis (c.F.) is exemplified by the altered concentrations of sodium and chloride in the sweat of c.F. subjects.’ In addition, sodium reabsorption is changed in rat salivary ducts perfused with sweat2 and saliva3 from c.F. patients. The salivary calcium level also changes in c.F. patients.4 Evidence of similar ionic changes promoted by sera of c.F, subjects has not been well documented. Sera from c.F. homozygotes have been extensively used, however, to demonstrate the presence of a ciliary dyskinesia factor (C.D.F.) which promotes mucociliary disturbances in cultured rabbit tracheal ciliated epithelial explants.5 C.D.F. is also found in sera from c.F. obligate heterozygotes and hence is thought to be related to
the defect in the c.F. disorder. A secretory response and an abnormality in
ciliary function, promoted in vitro by c.F.-serum C.D.F., have been experimentally induced in other mammalian’ both
W. PETTINGER T. PARKER A. HULL D. LONG R. PRATI
Pharmacology,
Departments of Pharmacology and Internal Medicine, University of Texas, Southwestern Medical School, Dallas, Texas 75235, U.S.A.
SALICYLATE INTERFERENCE WITH PLASMA-PARACETAMOL METHOD
SIR,-Plasma from patients with aspirin poisoning gives
a
reaction with the method for estimating paracetamol described by Glynn and Kendal.The concentration of salicylate in the plasma of patients admitted because of aspirin overdose was estimated by the method of Trinder.’ 16 samples (from nine patients) in which no paracetamol was detected by the gas-chromatographic method of Stewart and Willis8 were analysed by the method of Glynn and Kendal. The relationship between salicylate and apparent paracetamol concentrations in these samples is given by the equation p==0-7s+7, where p is the apparent paracetamol concentration in jg/ml and s is the salicylate concentration (as salicylic acid) in mg/dl. A closely similar relationship was found with aqueous solutions of sodium salicylate and with samples of salicylate-free plasma to which sodium salicylate was added to give concentrations up to 100 mg/d. Acetylsalicylic acid gave no reaction at a concentration of 100 mg/dl. Fortunately, the effect of salicylate on the method of Glynn and Kendal is not so great as to be likely to affect clinical decisions. Plasma containing 50 mg/dl salicylate would be expected to give a spurious value for paracetamol of 42 g/ml. The importance of the effect is that in interpreting results given by this method, it should be recognised that patients who have taken an overdose of aspirin have not necessarily also taken
false-positive
paracetamol. The magnitude of interference by salicylate may be reduced by modifying the method of Glynn and Kendal so that the absorbance of the final solution is measured at 450 nm instead of at 430 nm. This reduces interference by approximately 60% with negligible effect on sensitivity to paracetamol. With this minor modification, we believe that the method remains suitable for use in routine clinical-chemistry laboratories. The blood-paracetamol kit marketed by Winthrop Laboratories is based on the method of Glynn and Kendal and also l1ives snurious results with salicvlate.
Department of Clinical Chemistry, Hospital, Nottingham NG1 6HA General
PETER F. K. MACE GEOFFREY WALKER
Gill, J R Jr, George, J M., Solomon, A, Bartter, F. C. New Engl. J. Med. 1964, 270, 1088. 5 McAllister, R G., Van Way, S. W., Dayani, K., Anderson, W. J., Temple, E., Michelakis, A. M., Coppage, W. W., Oates, J A. Circulation Res.
4.
1971, 28/29, suppl
II,
12, 4.
to
those
as well as mollusc’ test systems. These cellular events were shown to be related to an influx of calcium ions from an extracellular source. In these test systems, such an influx of calcium was due to an increase in membrane permeability induced by calcium ionophore A23187. In an effort to determine whether increased permeability to calcium ions could explain the c.F.-serum C.D.F. response in the tracheal test system, we selectively increased the membrane permeability of the tracheal epithelium to calcium ions by adding calcium ionophore A23187 to the tracheal culture medium. We observed that the calcium ionophore A23187 produced the C.D.F.-like response only in the presence of a calcium level equivalent to that found in sera (2-25 to 2.75 mmol/1). Ionophore was also added to known normal control sera and the sera reassessed by bioassay. In addition, we chelated calcium in c.F. sera and culture medium with E.G.T.A. (ethyleneglycol-bis [beta-aminoethyl ether] N,N’ tetra-acetic acid) before bioassay, to study the effect of a calcium reduction on the tracheal-explant system.8 These studies determined that calcium ionophore A23187 in culture medium produced a change in mucociliary function in the tracheal test system indistinguishable from that promoted by c.F. sera. This c.D.F.-like response was shown to be directly proportional to the concentration of either calcium or ionophore. In addition, calcium chelation of culture medium with E.G.T.A. before bioassay abolished the ionophore-induced c.D.F.-like response. Addition of ionophore to normal control sera also induced the c.D.F.-like response, whereas E.G.T.A.-treated c.F. sera showed no c.D.F. activity by bioas-
say.8 These investigations, which are now being extended to include the ultrastructural alterations induced in the epithelium of the rabbit tracheal bioassay system, point to an important role for calcium as a cofactor in the production of the c.D.F. response generated by c.F. sera. The c.F. serum substance seems to act at the cellular-membrane level, promoting membrane-permeability changes (at least with respect to calcium) in the rabbit tracheal test system. Such a mechanism would not necessarily result in a net physiological imbalance of calcium in the rabbit tracheal system but only local changes in calcium concentration that would subsequently alter secretion and microtubular function. Since c.F. serum, like sweat’ and saliva3 from these same patients, can promote ionic fluxes, perhaps there is a common factor in all these body-fluids which 1. di 2. 3. 4.
5.
p. 160.
6. Glynn, J P., Kendal, S. E. Lancet, 1975, i, 1147. 7. Trinder, P Biochem. J. 1954, 57, 301. 8. Stewart, M. J., Willis, R G Ann. clin. Biochem. 1975,
comparable
6 7. 8
Sant ’Agnese, P., Darling, R., Perer, G., Shea, E. Pediatrics, Springfield. 1953, 12, 549 Mangos, J, McSherry, N., Benke, P. Pediat. Res. 1967, 1, 436 Mangos, J., McSherry, N. Science, 1967, 158, 135. Wotman, S., Mandel, I., Mercadante, J, Denning, C. R. Archs oral Biol 1971, 16, 663. Conover, J., Bonforte, R., Hathaway, P., Paciuc, S., Conod, E., Hirschhorn. K., Kopel, F. Pediat. Res. 1973, 7, 220. Cochrane, D., Douglas, W. Proc. natn. Acad. Sci. 1974, 71, 408. Satir, P Science, 1975, 190, 586. Bogart, B., Conod, E., Gaerlan, P., Conover, J. Pediat. Res. (in the press
by Gill et aB.4 and McAllister et al/ High plasmarenin-activity appeared to respond to control of accelerated hypertension by agents which have very little or no direct effect on renin release per se. Thus, the high blood-pressure could very well be the cause, and the raised renin activity the result, rather than vice versa as suggested by Dr Moore and demonstrated
Dr Goodwin. Division of Clinical
CALCIUM FLUX AND CYSTIC FIBROSIS
SiR,—That ionic disturbances are an important factor in cystic fibrosis (c.F.) is exemplified by the altered concentrations of sodium and chloride in the sweat of c.F. subjects.’ In addition, sodium reabsorption is changed in rat salivary ducts perfused with sweat2 and saliva3 from c.F. patients. The salivary calcium level also changes in c.F. patients.4 Evidence of similar ionic changes promoted by sera of c.F, subjects has not been well documented. Sera from c.F. homozygotes have been extensively used, however, to demonstrate the presence of a ciliary dyskinesia factor (C.D.F.) which promotes mucociliary disturbances in cultured rabbit tracheal ciliated epithelial explants.5 C.D.F. is also found in sera from c.F. obligate heterozygotes and hence is thought to be related to
the defect in the c.F. disorder. A secretory response and an abnormality in
ciliary function, promoted in vitro by c.F.-serum C.D.F., have been experimentally induced in other mammalian’ both
W. PETTINGER T. PARKER A. HULL D. LONG R. PRATI
Pharmacology,
Departments of Pharmacology and Internal Medicine, University of Texas, Southwestern Medical School, Dallas, Texas 75235, U.S.A.
SALICYLATE INTERFERENCE WITH PLASMA-PARACETAMOL METHOD
SIR,-Plasma from patients with aspirin poisoning gives
a
reaction with the method for estimating paracetamol described by Glynn and Kendal.The concentration of salicylate in the plasma of patients admitted because of aspirin overdose was estimated by the method of Trinder.’ 16 samples (from nine patients) in which no paracetamol was detected by the gas-chromatographic method of Stewart and Willis8 were analysed by the method of Glynn and Kendal. The relationship between salicylate and apparent paracetamol concentrations in these samples is given by the equation p==0-7s+7, where p is the apparent paracetamol concentration in jg/ml and s is the salicylate concentration (as salicylic acid) in mg/dl. A closely similar relationship was found with aqueous solutions of sodium salicylate and with samples of salicylate-free plasma to which sodium salicylate was added to give concentrations up to 100 mg/d. Acetylsalicylic acid gave no reaction at a concentration of 100 mg/dl. Fortunately, the effect of salicylate on the method of Glynn and Kendal is not so great as to be likely to affect clinical decisions. Plasma containing 50 mg/dl salicylate would be expected to give a spurious value for paracetamol of 42 g/ml. The importance of the effect is that in interpreting results given by this method, it should be recognised that patients who have taken an overdose of aspirin have not necessarily also taken
false-positive
paracetamol. The magnitude of interference by salicylate may be reduced by modifying the method of Glynn and Kendal so that the absorbance of the final solution is measured at 450 nm instead of at 430 nm. This reduces interference by approximately 60% with negligible effect on sensitivity to paracetamol. With this minor modification, we believe that the method remains suitable for use in routine clinical-chemistry laboratories. The blood-paracetamol kit marketed by Winthrop Laboratories is based on the method of Glynn and Kendal and also l1ives snurious results with salicvlate.
Department of Clinical Chemistry, Hospital, Nottingham NG1 6HA General
PETER F. K. MACE GEOFFREY WALKER
Gill, J R Jr, George, J M., Solomon, A, Bartter, F. C. New Engl. J. Med. 1964, 270, 1088. 5 McAllister, R G., Van Way, S. W., Dayani, K., Anderson, W. J., Temple, E., Michelakis, A. M., Coppage, W. W., Oates, J A. Circulation Res.
4.
1971, 28/29, suppl
II,
12, 4.
to
those
as well as mollusc’ test systems. These cellular events were shown to be related to an influx of calcium ions from an extracellular source. In these test systems, such an influx of calcium was due to an increase in membrane permeability induced by calcium ionophore A23187. In an effort to determine whether increased permeability to calcium ions could explain the c.F.-serum C.D.F. response in the tracheal test system, we selectively increased the membrane permeability of the tracheal epithelium to calcium ions by adding calcium ionophore A23187 to the tracheal culture medium. We observed that the calcium ionophore A23187 produced the C.D.F.-like response only in the presence of a calcium level equivalent to that found in sera (2-25 to 2.75 mmol/1). Ionophore was also added to known normal control sera and the sera reassessed by bioassay. In addition, we chelated calcium in c.F. sera and culture medium with E.G.T.A. (ethyleneglycol-bis [beta-aminoethyl ether] N,N’ tetra-acetic acid) before bioassay, to study the effect of a calcium reduction on the tracheal-explant system.8 These studies determined that calcium ionophore A23187 in culture medium produced a change in mucociliary function in the tracheal test system indistinguishable from that promoted by c.F. sera. This c.D.F.-like response was shown to be directly proportional to the concentration of either calcium or ionophore. In addition, calcium chelation of culture medium with E.G.T.A. before bioassay abolished the ionophore-induced c.D.F.-like response. Addition of ionophore to normal control sera also induced the c.D.F.-like response, whereas E.G.T.A.-treated c.F. sera showed no c.D.F. activity by bioas-
say.8 These investigations, which are now being extended to include the ultrastructural alterations induced in the epithelium of the rabbit tracheal bioassay system, point to an important role for calcium as a cofactor in the production of the c.D.F. response generated by c.F. sera. The c.F. serum substance seems to act at the cellular-membrane level, promoting membrane-permeability changes (at least with respect to calcium) in the rabbit tracheal test system. Such a mechanism would not necessarily result in a net physiological imbalance of calcium in the rabbit tracheal system but only local changes in calcium concentration that would subsequently alter secretion and microtubular function. Since c.F. serum, like sweat’ and saliva3 from these same patients, can promote ionic fluxes, perhaps there is a common factor in all these body-fluids which 1. di 2. 3. 4.
5.
p. 160.
6. Glynn, J P., Kendal, S. E. Lancet, 1975, i, 1147. 7. Trinder, P Biochem. J. 1954, 57, 301. 8. Stewart, M. J., Willis, R G Ann. clin. Biochem. 1975,
comparable
6 7. 8
Sant ’Agnese, P., Darling, R., Perer, G., Shea, E. Pediatrics, Springfield. 1953, 12, 549 Mangos, J, McSherry, N., Benke, P. Pediat. Res. 1967, 1, 436 Mangos, J., McSherry, N. Science, 1967, 158, 135. Wotman, S., Mandel, I., Mercadante, J, Denning, C. R. Archs oral Biol 1971, 16, 663. Conover, J., Bonforte, R., Hathaway, P., Paciuc, S., Conod, E., Hirschhorn. K., Kopel, F. Pediat. Res. 1973, 7, 220. Cochrane, D., Douglas, W. Proc. natn. Acad. Sci. 1974, 71, 408. Satir, P Science, 1975, 190, 586. Bogart, B., Conod, E., Gaerlan, P., Conover, J. Pediat. Res. (in the press