276 suffered may be attributable to this drug. We report this case as an example of the toxic effects which have occurred with the use of continual modest dosage. The drug has been withdrawn for four months now, but there seems little change in either the ocular or cutaneous manifestations. Connaught Hospital,
R. K. MALLYA M. D. R. MORRIS.
Walthamstow, London E17 9LW.
add a further possible case of practolol-induced peritoneal disease to the three reported by Dr Brown and his colleagues (Dec. 21, p. 1477). A woman of 58 had a six months’ history of abdominal distension and an acute episode of pain and vomiting. A firm mass Barium meal showed was palpable in the lower abdomen. dilatation of the stomach and duodenal loop, possibly due to adhesion at the duodenojejunal flexure. The remaining loops of small bowel appeared adherent to each other. Lymphangiogram and intravenous pyelogram were normal. At laparotomy a sheet of thickened peritoneum covered all the viscera and pelvic brim. There were adhesions between loops of bowel, and the palpable mass was thought to have been due to dilated loops of large bowel. The appearances suggested a chronic peritonitis, possibly tuberculous. Biopsy of the peritoneum showed a mixed acute and chronic inflammatory cell infiltrate without evidence of tuberculosis, the appearances being similar to that in the second case reported by Dr Brown and his colleagues (fig. 4, p. 1479). Cultures of peritoneum and peritoneal
SIR,-We should like
to
fluid were negative. There was no history of abdominal disease or surgery. She had been taking practolol (200 mg. daily) for four years, in conjunction with digoxin and frusemide for poorly controlled atrial fibrillation and heart-failure due to mitral-valve disease. Of particular interest was the associated development of keratoconjunctivitis sicca, a psoriasiform rash,l.2and a positive anti-nuclear factor test2 (titre 1/20) while taking practolol. The presence of these previously documented adverse reactions lends further support to the postulated link between practolol and peritoneal disease. -
Medical Unit, London Hospital, Whitechapel, London E1 1BB.
M. MINTON A. NEWLAND G. KNOWLES A. TURNBULL.
GENE DOSAGE EFFECT IN TRISOMY 21 SIR,-We read with interest your editorial (Dec. 28, p. 1554) on trisomy 21. We do not agree that no gene has been assigned to chromosome 21 and that no correlation between gene dosage and the activity of gene products has been established. Tan et awl. demonstrated that antiviral protein gene and indophenol oxidase gene (superoxide dismutase) (EC. 1. 15.1.1.) were both sited on chromosome 21. We have demonstrated4 that superoxide-dismutase activity (S.O.D.A.) is increased in the erythrocytes of trisomic-21 children. Their S.O.D.A. is found to be 1-4 times higher than for normal children. We think this is the first demonstration of a gene dosage effect in a trisomic condition. Likewise, Tan et awl. reported that trisomic-21 fibroblasts are 3-7 times more sensitive to protection by human interferon than normal diploid cells. These observations open a new field for research. P.-M. SINET D. ALLARD Hôpital des Enfants Malades, 149 rue de Sèvres, Paris 15e.
J. LEJEUNE H. JEROME.
Downie, C. C. Letter from I.C.I. Felix, R. H., Ive, F. A., Dahl, M. G. C. Br. med. J. 1974, iv, 321. Tan, Y. H., Tischfield, J., Ruddle, F. M. J. exp. Med. 1973, 137, 317. 4. Sinet, P.-M., Allard, D., Lejeune, J., Jerome, H. C. r. Acad. Sci. Paris, 1974, 278, 3267. 5. Tan, Y. H., Schneider, E. L., Tischfield, J., Epstein, C. J., Ruddle, F. H. Science, 1974, 186, 61. 1. 2. 3.
PREDNISONE IN DUCHENNE MUSCULAR DYSTROPHY
SiR,—The history of Duchenne muscular dystrophy (D.M.D.) is replete with therapeutic claims, all of which on more careful clinical analysis have been invalidated. Each new report of benefit is followed by another wave of disillusion among the families of patients and by an unnecessary effort by the scientific community to correct the impressions conveyed by well-meaning investigators. This is not to suggest that we must be so critical as to delay the application of any truly effective treatment. However, when the same mistake is made repeatedly, it behoves subsequent investigators to insist on unequivocally meaningful and clear-cut evidence. We fear that the report on prednisone in D.M.D. by Professor Drachman and his co-workers (Dec. 14, p. 1409) will prove another source of disappointment. They seem In our opinion they unaware of previous relevant studies. have presented a poorly constructed clinical investigation and conclusions unjustified by the data. The fact that this study was carried out by a group of workers whose academic standing is deservedly high increases our concern. Scattered reports of the efficacy of steroids in D.M.D. began to appear in the 1950s, but Dowben1 was the first to claim significant benefit. He claimed that only 8% of 37 patients with various forms of progressive muscular dystrophy continued to deteriorate while taking the anabolic steroid, 1-methyl-delta-androstenolone, either alone or in combination with digitoxin. Within the next least five studies 2-6 demonstrated lack of benefit from anabolic steroids. Unreported trials by eight other centres treating about 300 patients with D.M.D. also failed to produce significant benefit.44 Siegel et al.,7 in a three-year, well-controlled, double-blind trial failed to demonstrate significant improvement with prednisolone.
two years,
however,
at
.
’
We agree that neuromuscular
dysfunction, especially in evaluate. Nonetheless this should not preclude efforts to standardise and measure as many indices of weakness as possible. Such an attempt was made in three of the studies 2,6,7 with considerable Until we have a reliable success, even in children. biochemical indicator of worsening or improvement, it will be necessary to improve available quantitative techniques. Certainly parents’ assessments and measurements of strength by crude clinical ratings are not indicators of any statistical reliability. We have all seen apparent improvement in D.M.D. treated with prednisone when a diagnosis of children, is difficult
to strenuous
polymyositis had mistakenly been made; and, especially between the ages of 5 and 7 years, untreated patients may seemingly improve spontaneously as the processes of normal growth and development temporarily outstrip deterioration due to the disease process. The reasons given by the authors for the omission of any control cannot be accepted. Controls are even more mandatory when studying the effects of a drug such as prednisone which has prominentside-effects such as euphoria and sense of well-being. We agree that a reliable double-blind study with this drug is made difficult by its
recognisable side-effects. Nonetheless, by using a singledose alternate-day schedule (as was eventually done) these At the very least, clinical assessments can be minimised. 1. Dowben, R. M. New Engl. J. Med. 1963, 268, 912. 2. Barwick, D. D., Walton, J. N., Newell, D. J. Neurology, 1963, 13, 12. 3. Gamstorp, I. Acta pœdiat. scand. 1964, 53, 570. 4. Charash, L. Pediatrics, 1965, 36, 402. 5. Heyck, H., Laudahn, G., Leiders, C. J., Müller-Stephann, H., Schmidt-Peter, P. Acta pœdiat. scand. 1965, 54, 205. 6. Fowler, W. M., Pearson, C. M., Egstrom, G. H., Gardner, G. W. New Engl. J. Med. 1965, 272, 875. 7. Siegel, I. M., Miller, J. E., Ray, R. D. Illinois med. J. 1974, 145, 32.
277 should be carried out by someone with little or no knowledge of the treatment programme. Even if we set aside these criticisms, we find that the results do not fully justify the conclusions and the implied advice that prednisone should become a recognised treatment in D.M.D. In 6 of the 14 patients reported, therapy had to be discontinued: in 2 because of side-effects, in 1 because of lack of benefit, and in 3 because the parents were dissatisfied with the " trouble and risk " involved. We conclude that this in itself represents a 43% failure-rate. Half the patients had been treated for one year or less, 1 for only three weeks. In our judgment two years should be the absolute minimum of observation in any investigation of a drug in D.M.D. We agree with the statements that " because of the partial nature of the improvement the results have been difficult to evaluate " and that " the euphoria or ’increased energy’ produced by the prednisone might explain the improved functional state". We do not accept that their results suggest that a larger-scale investigation should be carried out. As Prof. George Schumacherhas emphasised for multiple-sclerosis research, experience suggests that largescale pharmacological trials in neurological disorders should be carried out only when an agent clearly demonstrates benefit in preliminary studies. Transient or equivocal benefit is not sufficient. Conclusions are doubly hazardous when objective measurements of clinical or biochemical improvement are lacking. We would encourage the authors to publish a follow-up report in 1-2 years, since we believe the apparent beneficial ,effects of treatment will not be sustained. Department of Neurology, University of Newcastle upon Tyne, THEODORE L. MUNSAT Newcastle General Hospital, Newcastle upon Tyne NE4 6BE. JOHN N. WALTON.
MESANGIAL AND KUPFFER CELLS
SIR,-Dr Bjorneboe’s hypothesis (Jan. 4, p. 45) on the Kupffer cell in cirrhosis is interesting in its possible analogy with the kidney. The function of the Kupffer cell would seem similar to that of the glomerular mesangial cell in that both may remove toxic materials before damage to the organ occurs. Most forms of glomerular disease appear to be immune-complex (antibodyplus-complement) associated,9 and it seems likely that in disease associated with removal of complexes by the mesangial cell-e.g., focal proliferative glomerulonephritis, IgA nephropathy, and diffuse exudative proliferative (G.N.)10 -the patient may have a good prognosis,11 and where complexes escape from the mesangial cell and deposit along the glomerular capillary wall-e.g., epimembranous, mesangiocapillary glomerulonephritis-the prognosis is poor." Further in the NZB/W hybrid mouse, which develops a spontaneous lupus-like nephropathy, immune deposits may be detected in the mesangium before any histological or clinical renal disease, while later on, when overt renal disease appears, the complexes are not seen in the mesangium but along the capillary wa11.12This suggests that when the mesangial cell fails to recognise or remove immune complexes the prognosis is worsened. That the failure may be more a failure of removal is perhaps suggested by mesangiocapillary glomerulonephritis (M.C.G.), where the mesangial cell may be seen to push processes role of the
8. 9.
Schumacher, G. A. Neurology, 1974, 24, 1010. Peters, D. K. in Clinical Aspects of Immunology (edited by P. G. H. Gell, R. R. A. Coombs, and P. J. Lachmann). Oxford, 1974. 10. Davison, A. M., Thomson, D., Macdonald, M. K., Uttley, W. S., Robson, J. S. J. clin. Path. 1973, 26, 198. 11. Cameron, J. S. Br. med. J. 1972, iv, 87. 12. Leathem, A. Unpublished.
round the glomerular capillary wall towards and covering endomembranous deposits but does not appear to engulf them, suggesting the mesangial cells can recognise complexes but are unable to remove them. The role of such factors as macrophage receptors and immune-complex composition is unknown in renal disease, but on size alone the endomembranous deposits seen in M.C.G. and diffuse lupus nephropathy should be removable from their relatively accessible positions. Such a failure may represent some blockage to the reticuloendothelial system or more particularly the mesangial cell and perhaps, as suggested by Dr Bjorneboe for patients with cirrhosis, renal patients with M.C.G. and diffuse proliferative lupus nephropathy might benefit from a trial of some reticuloendothelial stimulant. Bland-Sutton Institute of
Pathology, Middlesex Hospital, London W1N 8AA.
A. LEATHEM.
COLISTIN SENSITIVITY TESTING SIR,-In reply to the letter by Dr Edmunds (Dec. 21, p. 1526) I should like to make the following points. ’ Colomycin ’ is indeed administered parenterally in the form of colistin sulphomethate sodium (c.s.M.s.). The activity of c.s.M.s. is 12,500 units per mg. while that of colistin sulphate (c.s.) is 19,500 units per mg., and that of colistin base is 30,000 units ’per mg. Unlike Eickhoff and Finland,l who showed that c.s. is about eight times more active than c.s.M.s., it is our experience that the activity of c.s. is approximately twice that of c.s.M.s. c.s.mt.s. is obtained by the sulphomethylation of the five free amino groups of colistin base. c.s.M.s. does tend to hydrolyse in the body with the progressive loss of the sulphomethyl groups.2 These compounds of intermediate substitution would probably have activities in units per mg. between c.s.M.s. and colistin sulphate. The determination of colomycin levels is exceptionally complex because one is dealing with an antibiotic that hydrolyses to a more active compound which has different molecular properties. serum level of colomycin is measured by comparing the of inhibition of an appropriate serum dilution with that produced by a standard of known concentration. If blood samples are taken about 1 to 3 hours after injection most of the colistin present will still be fairly highly substituted; therefore, the nearest standard would be C.S.M.S.2 Because of their different molecular properties, which results in a different degree of diffusion through agar, the concentration of a solution containing c.s.M.s. cannot be obtained by comparing the zone of inhibition with the zone of inhibition produced with c.s. The different structures of colistin base, C.S.M.S. and c.s., are responsible for the varying activities. If the serum contains compounds which are less highly substituted than c.s.M.s., these will tend to be more active and to diffuse faster through the agar producing a slightly larger zone of inhibition and a somewhat higher serum level than is actually present. Once a serum concentration of colomycin relative to c.s.M.s. has been obtained this can be converted to units and to the
The
zone
equivalent concentration of c.s. Alternatively, it would be preferable to express all results in terms of units. We now routinely employ 25 µg. and 100 µg. colistin sulphate discs for sensitivity testing since this compound (1) does not produce the occasional misleading result indicating sensitivity of staphylococci which occurred with c.s.M.s., (2) is cheaper, (3) is more stable, (4) diffuses more easily, providing a clearer zone of inhibition. (Colomycin diffuses poorly in agar, and it is necessary allow the antibiotic before incubation.)
to
1. 2.
to
diffuse at
room
temperature for 3 hours
Eickhoff, T. C., Finland, M. Am. J. med. Sci. 1965, 249, 172. Beveridge, G. G., Martin, A. J. Br. J. Pharmac. Chemother. 1967, 29, 125.
R. K. MALLYA M. D. R. MORRIS.
Walthamstow, London E17 9LW.
add a further possible case of practolol-induced peritoneal disease to the three reported by Dr Brown and his colleagues (Dec. 21, p. 1477). A woman of 58 had a six months’ history of abdominal distension and an acute episode of pain and vomiting. A firm mass Barium meal showed was palpable in the lower abdomen. dilatation of the stomach and duodenal loop, possibly due to adhesion at the duodenojejunal flexure. The remaining loops of small bowel appeared adherent to each other. Lymphangiogram and intravenous pyelogram were normal. At laparotomy a sheet of thickened peritoneum covered all the viscera and pelvic brim. There were adhesions between loops of bowel, and the palpable mass was thought to have been due to dilated loops of large bowel. The appearances suggested a chronic peritonitis, possibly tuberculous. Biopsy of the peritoneum showed a mixed acute and chronic inflammatory cell infiltrate without evidence of tuberculosis, the appearances being similar to that in the second case reported by Dr Brown and his colleagues (fig. 4, p. 1479). Cultures of peritoneum and peritoneal
SIR,-We should like
to
fluid were negative. There was no history of abdominal disease or surgery. She had been taking practolol (200 mg. daily) for four years, in conjunction with digoxin and frusemide for poorly controlled atrial fibrillation and heart-failure due to mitral-valve disease. Of particular interest was the associated development of keratoconjunctivitis sicca, a psoriasiform rash,l.2and a positive anti-nuclear factor test2 (titre 1/20) while taking practolol. The presence of these previously documented adverse reactions lends further support to the postulated link between practolol and peritoneal disease. -
Medical Unit, London Hospital, Whitechapel, London E1 1BB.
M. MINTON A. NEWLAND G. KNOWLES A. TURNBULL.
GENE DOSAGE EFFECT IN TRISOMY 21 SIR,-We read with interest your editorial (Dec. 28, p. 1554) on trisomy 21. We do not agree that no gene has been assigned to chromosome 21 and that no correlation between gene dosage and the activity of gene products has been established. Tan et awl. demonstrated that antiviral protein gene and indophenol oxidase gene (superoxide dismutase) (EC. 1. 15.1.1.) were both sited on chromosome 21. We have demonstrated4 that superoxide-dismutase activity (S.O.D.A.) is increased in the erythrocytes of trisomic-21 children. Their S.O.D.A. is found to be 1-4 times higher than for normal children. We think this is the first demonstration of a gene dosage effect in a trisomic condition. Likewise, Tan et awl. reported that trisomic-21 fibroblasts are 3-7 times more sensitive to protection by human interferon than normal diploid cells. These observations open a new field for research. P.-M. SINET D. ALLARD Hôpital des Enfants Malades, 149 rue de Sèvres, Paris 15e.
J. LEJEUNE H. JEROME.
Downie, C. C. Letter from I.C.I. Felix, R. H., Ive, F. A., Dahl, M. G. C. Br. med. J. 1974, iv, 321. Tan, Y. H., Tischfield, J., Ruddle, F. M. J. exp. Med. 1973, 137, 317. 4. Sinet, P.-M., Allard, D., Lejeune, J., Jerome, H. C. r. Acad. Sci. Paris, 1974, 278, 3267. 5. Tan, Y. H., Schneider, E. L., Tischfield, J., Epstein, C. J., Ruddle, F. H. Science, 1974, 186, 61. 1. 2. 3.
PREDNISONE IN DUCHENNE MUSCULAR DYSTROPHY
SiR,—The history of Duchenne muscular dystrophy (D.M.D.) is replete with therapeutic claims, all of which on more careful clinical analysis have been invalidated. Each new report of benefit is followed by another wave of disillusion among the families of patients and by an unnecessary effort by the scientific community to correct the impressions conveyed by well-meaning investigators. This is not to suggest that we must be so critical as to delay the application of any truly effective treatment. However, when the same mistake is made repeatedly, it behoves subsequent investigators to insist on unequivocally meaningful and clear-cut evidence. We fear that the report on prednisone in D.M.D. by Professor Drachman and his co-workers (Dec. 14, p. 1409) will prove another source of disappointment. They seem In our opinion they unaware of previous relevant studies. have presented a poorly constructed clinical investigation and conclusions unjustified by the data. The fact that this study was carried out by a group of workers whose academic standing is deservedly high increases our concern. Scattered reports of the efficacy of steroids in D.M.D. began to appear in the 1950s, but Dowben1 was the first to claim significant benefit. He claimed that only 8% of 37 patients with various forms of progressive muscular dystrophy continued to deteriorate while taking the anabolic steroid, 1-methyl-delta-androstenolone, either alone or in combination with digitoxin. Within the next least five studies 2-6 demonstrated lack of benefit from anabolic steroids. Unreported trials by eight other centres treating about 300 patients with D.M.D. also failed to produce significant benefit.44 Siegel et al.,7 in a three-year, well-controlled, double-blind trial failed to demonstrate significant improvement with prednisolone.
two years,
however,
at
.
’
We agree that neuromuscular
dysfunction, especially in evaluate. Nonetheless this should not preclude efforts to standardise and measure as many indices of weakness as possible. Such an attempt was made in three of the studies 2,6,7 with considerable Until we have a reliable success, even in children. biochemical indicator of worsening or improvement, it will be necessary to improve available quantitative techniques. Certainly parents’ assessments and measurements of strength by crude clinical ratings are not indicators of any statistical reliability. We have all seen apparent improvement in D.M.D. treated with prednisone when a diagnosis of children, is difficult
to strenuous
polymyositis had mistakenly been made; and, especially between the ages of 5 and 7 years, untreated patients may seemingly improve spontaneously as the processes of normal growth and development temporarily outstrip deterioration due to the disease process. The reasons given by the authors for the omission of any control cannot be accepted. Controls are even more mandatory when studying the effects of a drug such as prednisone which has prominentside-effects such as euphoria and sense of well-being. We agree that a reliable double-blind study with this drug is made difficult by its
recognisable side-effects. Nonetheless, by using a singledose alternate-day schedule (as was eventually done) these At the very least, clinical assessments can be minimised. 1. Dowben, R. M. New Engl. J. Med. 1963, 268, 912. 2. Barwick, D. D., Walton, J. N., Newell, D. J. Neurology, 1963, 13, 12. 3. Gamstorp, I. Acta pœdiat. scand. 1964, 53, 570. 4. Charash, L. Pediatrics, 1965, 36, 402. 5. Heyck, H., Laudahn, G., Leiders, C. J., Müller-Stephann, H., Schmidt-Peter, P. Acta pœdiat. scand. 1965, 54, 205. 6. Fowler, W. M., Pearson, C. M., Egstrom, G. H., Gardner, G. W. New Engl. J. Med. 1965, 272, 875. 7. Siegel, I. M., Miller, J. E., Ray, R. D. Illinois med. J. 1974, 145, 32.
277 should be carried out by someone with little or no knowledge of the treatment programme. Even if we set aside these criticisms, we find that the results do not fully justify the conclusions and the implied advice that prednisone should become a recognised treatment in D.M.D. In 6 of the 14 patients reported, therapy had to be discontinued: in 2 because of side-effects, in 1 because of lack of benefit, and in 3 because the parents were dissatisfied with the " trouble and risk " involved. We conclude that this in itself represents a 43% failure-rate. Half the patients had been treated for one year or less, 1 for only three weeks. In our judgment two years should be the absolute minimum of observation in any investigation of a drug in D.M.D. We agree with the statements that " because of the partial nature of the improvement the results have been difficult to evaluate " and that " the euphoria or ’increased energy’ produced by the prednisone might explain the improved functional state". We do not accept that their results suggest that a larger-scale investigation should be carried out. As Prof. George Schumacherhas emphasised for multiple-sclerosis research, experience suggests that largescale pharmacological trials in neurological disorders should be carried out only when an agent clearly demonstrates benefit in preliminary studies. Transient or equivocal benefit is not sufficient. Conclusions are doubly hazardous when objective measurements of clinical or biochemical improvement are lacking. We would encourage the authors to publish a follow-up report in 1-2 years, since we believe the apparent beneficial ,effects of treatment will not be sustained. Department of Neurology, University of Newcastle upon Tyne, THEODORE L. MUNSAT Newcastle General Hospital, Newcastle upon Tyne NE4 6BE. JOHN N. WALTON.
MESANGIAL AND KUPFFER CELLS
SIR,-Dr Bjorneboe’s hypothesis (Jan. 4, p. 45) on the Kupffer cell in cirrhosis is interesting in its possible analogy with the kidney. The function of the Kupffer cell would seem similar to that of the glomerular mesangial cell in that both may remove toxic materials before damage to the organ occurs. Most forms of glomerular disease appear to be immune-complex (antibodyplus-complement) associated,9 and it seems likely that in disease associated with removal of complexes by the mesangial cell-e.g., focal proliferative glomerulonephritis, IgA nephropathy, and diffuse exudative proliferative (G.N.)10 -the patient may have a good prognosis,11 and where complexes escape from the mesangial cell and deposit along the glomerular capillary wall-e.g., epimembranous, mesangiocapillary glomerulonephritis-the prognosis is poor." Further in the NZB/W hybrid mouse, which develops a spontaneous lupus-like nephropathy, immune deposits may be detected in the mesangium before any histological or clinical renal disease, while later on, when overt renal disease appears, the complexes are not seen in the mesangium but along the capillary wa11.12This suggests that when the mesangial cell fails to recognise or remove immune complexes the prognosis is worsened. That the failure may be more a failure of removal is perhaps suggested by mesangiocapillary glomerulonephritis (M.C.G.), where the mesangial cell may be seen to push processes role of the
8. 9.
Schumacher, G. A. Neurology, 1974, 24, 1010. Peters, D. K. in Clinical Aspects of Immunology (edited by P. G. H. Gell, R. R. A. Coombs, and P. J. Lachmann). Oxford, 1974. 10. Davison, A. M., Thomson, D., Macdonald, M. K., Uttley, W. S., Robson, J. S. J. clin. Path. 1973, 26, 198. 11. Cameron, J. S. Br. med. J. 1972, iv, 87. 12. Leathem, A. Unpublished.
round the glomerular capillary wall towards and covering endomembranous deposits but does not appear to engulf them, suggesting the mesangial cells can recognise complexes but are unable to remove them. The role of such factors as macrophage receptors and immune-complex composition is unknown in renal disease, but on size alone the endomembranous deposits seen in M.C.G. and diffuse lupus nephropathy should be removable from their relatively accessible positions. Such a failure may represent some blockage to the reticuloendothelial system or more particularly the mesangial cell and perhaps, as suggested by Dr Bjorneboe for patients with cirrhosis, renal patients with M.C.G. and diffuse proliferative lupus nephropathy might benefit from a trial of some reticuloendothelial stimulant. Bland-Sutton Institute of
Pathology, Middlesex Hospital, London W1N 8AA.
A. LEATHEM.
COLISTIN SENSITIVITY TESTING SIR,-In reply to the letter by Dr Edmunds (Dec. 21, p. 1526) I should like to make the following points. ’ Colomycin ’ is indeed administered parenterally in the form of colistin sulphomethate sodium (c.s.M.s.). The activity of c.s.M.s. is 12,500 units per mg. while that of colistin sulphate (c.s.) is 19,500 units per mg., and that of colistin base is 30,000 units ’per mg. Unlike Eickhoff and Finland,l who showed that c.s. is about eight times more active than c.s.M.s., it is our experience that the activity of c.s. is approximately twice that of c.s.M.s. c.s.mt.s. is obtained by the sulphomethylation of the five free amino groups of colistin base. c.s.M.s. does tend to hydrolyse in the body with the progressive loss of the sulphomethyl groups.2 These compounds of intermediate substitution would probably have activities in units per mg. between c.s.M.s. and colistin sulphate. The determination of colomycin levels is exceptionally complex because one is dealing with an antibiotic that hydrolyses to a more active compound which has different molecular properties. serum level of colomycin is measured by comparing the of inhibition of an appropriate serum dilution with that produced by a standard of known concentration. If blood samples are taken about 1 to 3 hours after injection most of the colistin present will still be fairly highly substituted; therefore, the nearest standard would be C.S.M.S.2 Because of their different molecular properties, which results in a different degree of diffusion through agar, the concentration of a solution containing c.s.M.s. cannot be obtained by comparing the zone of inhibition with the zone of inhibition produced with c.s. The different structures of colistin base, C.S.M.S. and c.s., are responsible for the varying activities. If the serum contains compounds which are less highly substituted than c.s.M.s., these will tend to be more active and to diffuse faster through the agar producing a slightly larger zone of inhibition and a somewhat higher serum level than is actually present. Once a serum concentration of colomycin relative to c.s.M.s. has been obtained this can be converted to units and to the
The
zone
equivalent concentration of c.s. Alternatively, it would be preferable to express all results in terms of units. We now routinely employ 25 µg. and 100 µg. colistin sulphate discs for sensitivity testing since this compound (1) does not produce the occasional misleading result indicating sensitivity of staphylococci which occurred with c.s.M.s., (2) is cheaper, (3) is more stable, (4) diffuses more easily, providing a clearer zone of inhibition. (Colomycin diffuses poorly in agar, and it is necessary allow the antibiotic before incubation.)
to
1. 2.
to
diffuse at
room
temperature for 3 hours
Eickhoff, T. C., Finland, M. Am. J. med. Sci. 1965, 249, 172. Beveridge, G. G., Martin, A. J. Br. J. Pharmac. Chemother. 1967, 29, 125.
