1104
fluidity, and the transport also be important.99
of
carcinogens
into the cells, may
With these modifications and precautions, the coefficient of variation for triplicate determinations was approximately 10% in a study of 41 cases of laryngeal carcinoma.lo Assessment of A.H.H. inducibility may be important in identifying individuals at risk of cancer associated with exposure to P.A.H. Department 1 and Department of Lung Medicine, University Hospital, and Department of Tumour Cytogenetics, Wallenberg Laboratory, University of Lund, S-220 07 Lund, Sweden Research
ROLF KORSGAARD
Section of Preventive Medicine, Department of Internal Medicine,
University of Lund, Malmö, Sweden
ERIK TRELL
TISSUE PENETRATION BY CEPHALOSPORINS
SIR,-Pharmacologists and medical microbiologists have long been preoccupied with serum concentrations of antibiotic, even though drug levels in tissue fluids, transudates, and exudates are far more relevant to the treatment of infection,’ so the study by Dr Gillett and Dr Wise (May 6, p. 962) is welcome. However, the conclusion "that the penetration of antibiotic into tissue fluid does not depend on the fraction of free drug in the blood" is not supported by their results. The cephamycin, cefoxitin, was the most highly proteinbound of the four "cephalosporins" tested, and was the only one that produced no detectable concentration in the transudate from the area of abraded skin two hours after drug adPHARMACOKINETIC PROPERTIES OF CEPHALOSPORINS
*Apparent volume of distribution (1/1-73 m2); cephalothin ficially high due to metabolism.
value arti-
ministration. Gillett and Wise explain this
on the basis of the half-life of cefoxitin, yet the cephalosporin CGP 9000, with the same short half-life, produced a substantial concentration in the abrasion fluid at three hours. As Gillett and Wise say, "what is not known is the level of free drug in the tissue fluid". Their antibiotic assay method did not distinguish between free drug and protein-bound drug. Even with a lower albumin content than that of plasma, the abrasion fluid would almost certainly contain a considerable proportion of bound cefoxitin and, to a lesser extent, cefuroxime, while both cephalexin and CGP 9000 would be present almost entirely as free antibiotic.2 Only free drug can exert antimicrobial activity.3 After reviewing their own extensive studies and other published work, Craig and Welling3 could find only one anomalous publication. This is the study by Waterman et a1.4 which Gil-
short
serum
9. Pero, R. Unpublished. 10. Korsgaard, R., and others. Unpublished. 1. Selwyn, S. Lancet, 1976, ii, 808. 2. Craig, W. A. Infection, 1976, 4, s160. 3. Craig, W. A., Welling, P. G. Clin. Pharmacokin. 1977, 2, 252. 4. Waterman, N. G., Raff, M. J., Scharfenberger, L., Barnwell, P. Dis. 1976, 133, 642.
A. J. infect.
lett and Wise cite in support of their viewpoint. However, Craig and Welling had no hesitation in deciding that "in spite of the array of different models and animal species, the ... conclusion to be drawn is that distribution and penetration of antimicrobial agents into these [extravascular] fluids is primarily dependent on the fraction of free drug [in the plasma]". In its turn the fraction of free drug in the tissue fluid should be regarded as the therapeutically relevant entity which requires our detailed attention. Gillett and Wise suggest that the longer the serum half-life of an antibiotic the higher may be its tissue concentration (although no such clear relationship is seen in their results). However, a drug cannot be in two places at once, so a long serum half-life indicates a relative inability of the drug to leave the bloodstream and enter other body compartments or be eliminated. The all-important tissue-fluid half-life of free drug is likely to be determined more by the initial peak serum concentration of free antibiotic than by the total-drug serum half-life, and is reflected in the "apparent volume of distribution of the drug". Pharmacokinetic values for the principal cephalosporins3are shown in the table. Department of Bacteriology, Westminster Medical School, London SW1P 2AR
***This letter has been shown whose reply follows.-ED. L.
SYDNEY SELWYN
to
Dr Wise and Dr
Gillett,
SIR,-Firstly, to answer Dr Selwyn’s comment on the lack of detectable cefoxitin in the tissue transudate at 2 z 3h. If all four drugs are compared it is immediately apparent that the tissue levels are closely related to the serum level (and, therefore, serum half-life). The 3 h level of cefoxitin would, therefore, by extrapolation, be 1-2 flg/ml which, as we state, is below the sensitivity of the assay. It is inappropriate to compare cephalexin and CGP 9000, given orally and having a peak serum concentration at 2 h (absorption continuing for some time thereafter), with intravenous cefoxitin. Serum halflives are calculated from the elimination phase. We agree that the free level of an antibiotic in the tissue fluid is of great relevance. We are studying this subject, using a more versatile blister technique to measure both free and bound drug. However, it is of considerable interest that high protein binding does not seem to affect the penetration and concentration of total antibiotic in the tissue. This is an important observation in its own right. Dr Selwyn is incorrect in thinking that "a long serum halflife indicates a relative inability of the drug to leave the bloodstream and enter other body compartments". A well-distributed drug is often more slowly eliminated because these "deeper" compartments act as a depot. Good examples of welldistributed drugs are trimethopriml and griseofulvin,2 with serum half-lives in excess of 9 h. The serum half-life may well (but not necessarily) be of importance in determining an antibiotic’s penetration into tissues. Department of Medical Microbiology, RICHARD WISE Dudley Road Hospital, A. P. GILLETT Birmingham B18 7QH
MUCOCUTANEOUS REACTIONS DURING MYCOPLASMA PNEUMONIÆ INFECTION
SIR,-Dr Lind (March 25, p. 655) found that, among patients with raised cold agglutinin titres, Stevens-Johnson syndrome and non-specific exanthemas were more common in those with positive Mycoplasma pneumoniae serology. 5. Gower, P. E., Dash, C. H. Eur. J. clin. Pharmac. 1977, 12, 221. 1. Fowle, A. S. E. in Trimethoprim/sulphamethoxazole in Bacterial Infections (edited by L. S. Bernstein and A. J. Salter); p. 63. Edinburgh, 1973. 2. Rowland, M., Reigelman, S., Epstein, W. L. J. pharm. Sci. 1968, 57, 984.
1105 We have studied all children referred to Bristol hospitals during an M. pneumonia epidemic lasting 18 months. 9 (20%) out of 44 children with serologically proven M. pneumoniae infection had rashes. 6 children had an erythematous macular rash on the trunk and limbs, and this was usually fading at the time of referral. In 1 of the 6 Stevens-Johnson syndrome developed and there were 3 additional cases of the syndrome. In 3 of the 4 cold agglutinins were sought and all 3 were positive. These findings do not clarify the role of cold agglutinins in the pathogenesis of mycoplasma-associated mucocutaneous reactions. 1 of the children with Stevens-Johnson syndrome, a girl of 11 years, had extensive toxic epidermal necrolysis with typical "scalded skin" erythema, blistering, and epidermal stripping. This rare complication caused significant fluid depletion, but the skin healed within 14 days with no further major problem. The conjunctival ulceration, however, caused healing and fibrosis of such intensity that severe conjunctival shrinkage syndrome, corneal scarring, and visual impairment ensued. 1 case of Stevens-Johnson syndrome was not associated with lower-respiratory-tract infection, which supports Lind’s view that M. pneumonia infection can cause rashes without clinical evidence of respiratory disease.
Cherry et al.2 found no published cases of Stevens-Johnson syndrome associated with M. pneumoniae in girls; 2 of our cases were
females.
Although patients with Stevens-Johnson syndrome usually recover fully we recommend that, in view of the possibility of permanent eye damage, all children with the syndrome should be referred to an ophthalmologist. Royal Devon and
Exeter
Hospital,
Exeter EX2 5DW
Gloucestershire Royal Gloucester GL1 3NN
MEASUREMENT OF LACTOFERRIN IN PANCREATIC JUICE described a protein in pancreatic juice of calcifying pancreatitis;’ this was later identified as lactoferrin,2 a glycoprotein mainly secreted by mammary glands and found in various external secretions.3 Indirect
SIR,-In 1971
men
we
with chronic
immunofluorescence studies revealed lactoferrin in acinar cells of normal pancreatic tissue and in acinar cells of men with chronic calcifying pancreatitis.4 However, the fluorescence was more striking in pathological material. Dr Fedail and colleagues (Jan.. 28, p. 181) have confirmed that lactoferrin secretion is increased in pancreatic juice of patients with chronic
pancreatitis. Since lactoferrin secretion is not increased in pancreatic in acute pancreatitis,s a practical and accurate assay for lactoferrin in pancreatic juice might be a useful diagnostic test. We have compared a double-diffusion technique with a cancer or
radioimmunoassay. The radioimmunoassay was done with pure human lactoferrin (kindly given by Dr P. Masson) labelled with 125by the chloramine-T method. Separation of bound and free lactoferrin was achieved by a double-antibody technique. The sensitivity of the assay was 1 ng/ml. The antilactoferrin prepared in the laboratory was used after a dilution of 1/50 000. ’Immunoplates LC’ (low-concentration) ’Partigen’ lactoferrin and pure standard lactoferrin (titrating 4 mg/dl) were kindly prepared by Dr Baudner (Behring Institute,. Marburg).
1.
PETER G. F. SWIFT
Hospital, DAVID W. STEVENS
Clemente, F., Ribeiro, T., Colomb, E., Figarella, C., Sarles, H. Biochim. biophys. Acta, 1971, 251, 456. 2. Colomb, E., Estevenon, J. P., Figarella, C., Guy, O., Sarles, H. ibid. 1974, 342, 306. 3. Masson, P. La lactoferrine. Paris, 1970. 4. Colomb, E., Pianetta, C., Estevenon, J. P., Guy, O., Figarella, C., Sarles, H. Digestion, 1976, 14, 242. 5. Estevenon, J. P., Sarles, H., Figarella, C. Scand. J. Gastroent. 1975, 10, 327.
SERRATIA MARCESCENS COLONISING THE GUT
SIR,-During an outbreak of hospital cross-infections by multiply resistant Serratia marcescens this organism was occasionally recovered from stools as well as from the site of clinical infection. The spread of Klebsiella pneumoniae from a gastrointestinal-tract reservoir to extraintestinal sites is well documented,’ but S. marcescens does not usually colonise the gut of adults.2 We extended our observation by taking rectal swabs from patients admitted during the outbreak; the swab was streaked on selective media and the plate was inspected caused
after 24 and 48 h incubation at 37°C; the isolates were identified and antibiotic susceptibilities were determined by standard methods. 120 rectal cultures were obtained from 57 patients. The stools of 5 patients were colonised with multiply resistant S. marcescens. 2 patients were colonised or infected at extraintestinal sites at the time the first stool culture was obtained; the other 3 patients later became colonised or infected at extraintestinal sites. As with K. pneumonice, gastrointestinal colonisation by S. marcescens may, by serving as a reservoir for cross-infection, be an important step in the development of hospital-acquired infection. Infectious Disease Section, Department of Medicine, Veterans Administration Hospital, University of Oregon Health Science Center, Portland, Oregon 97207, U.S.A.
STEPHEN R. JONES MERIDITH AMON CATHERINE FALVEY KATHRYN PATRICK
Comparative 1.
Stevens, D, Swift, P. G. F., Johnston, P. G. B., Kearney, P. J., Corner, B. D., Burman, D. Archs Dis. Childh. 1978, 53, 38. 2 Cherry, J. D., Hurwitz, E. S., Welliver, R. C. J. Pediat. 1975, 87, 369. 1. Selden, R., and others Ann. intern. Med. 1971, 74, 6737. 2. Farmer, J. J, and others Lancet, 1976, ii, 455.
measurement of lactoferrin.
Radioimmunoassay (e);
assay
by
radial immunodiffusion
(0).
Abs-
cissa ; pancreatic juice from patients with chronic pancreatitis (1-6) and controls values.
(7-12).
The dotted line represents the limit of normal
fluidity, and the transport also be important.99
of
carcinogens
into the cells, may
With these modifications and precautions, the coefficient of variation for triplicate determinations was approximately 10% in a study of 41 cases of laryngeal carcinoma.lo Assessment of A.H.H. inducibility may be important in identifying individuals at risk of cancer associated with exposure to P.A.H. Department 1 and Department of Lung Medicine, University Hospital, and Department of Tumour Cytogenetics, Wallenberg Laboratory, University of Lund, S-220 07 Lund, Sweden Research
ROLF KORSGAARD
Section of Preventive Medicine, Department of Internal Medicine,
University of Lund, Malmö, Sweden
ERIK TRELL
TISSUE PENETRATION BY CEPHALOSPORINS
SIR,-Pharmacologists and medical microbiologists have long been preoccupied with serum concentrations of antibiotic, even though drug levels in tissue fluids, transudates, and exudates are far more relevant to the treatment of infection,’ so the study by Dr Gillett and Dr Wise (May 6, p. 962) is welcome. However, the conclusion "that the penetration of antibiotic into tissue fluid does not depend on the fraction of free drug in the blood" is not supported by their results. The cephamycin, cefoxitin, was the most highly proteinbound of the four "cephalosporins" tested, and was the only one that produced no detectable concentration in the transudate from the area of abraded skin two hours after drug adPHARMACOKINETIC PROPERTIES OF CEPHALOSPORINS
*Apparent volume of distribution (1/1-73 m2); cephalothin ficially high due to metabolism.
value arti-
ministration. Gillett and Wise explain this
on the basis of the half-life of cefoxitin, yet the cephalosporin CGP 9000, with the same short half-life, produced a substantial concentration in the abrasion fluid at three hours. As Gillett and Wise say, "what is not known is the level of free drug in the tissue fluid". Their antibiotic assay method did not distinguish between free drug and protein-bound drug. Even with a lower albumin content than that of plasma, the abrasion fluid would almost certainly contain a considerable proportion of bound cefoxitin and, to a lesser extent, cefuroxime, while both cephalexin and CGP 9000 would be present almost entirely as free antibiotic.2 Only free drug can exert antimicrobial activity.3 After reviewing their own extensive studies and other published work, Craig and Welling3 could find only one anomalous publication. This is the study by Waterman et a1.4 which Gil-
short
serum
9. Pero, R. Unpublished. 10. Korsgaard, R., and others. Unpublished. 1. Selwyn, S. Lancet, 1976, ii, 808. 2. Craig, W. A. Infection, 1976, 4, s160. 3. Craig, W. A., Welling, P. G. Clin. Pharmacokin. 1977, 2, 252. 4. Waterman, N. G., Raff, M. J., Scharfenberger, L., Barnwell, P. Dis. 1976, 133, 642.
A. J. infect.
lett and Wise cite in support of their viewpoint. However, Craig and Welling had no hesitation in deciding that "in spite of the array of different models and animal species, the ... conclusion to be drawn is that distribution and penetration of antimicrobial agents into these [extravascular] fluids is primarily dependent on the fraction of free drug [in the plasma]". In its turn the fraction of free drug in the tissue fluid should be regarded as the therapeutically relevant entity which requires our detailed attention. Gillett and Wise suggest that the longer the serum half-life of an antibiotic the higher may be its tissue concentration (although no such clear relationship is seen in their results). However, a drug cannot be in two places at once, so a long serum half-life indicates a relative inability of the drug to leave the bloodstream and enter other body compartments or be eliminated. The all-important tissue-fluid half-life of free drug is likely to be determined more by the initial peak serum concentration of free antibiotic than by the total-drug serum half-life, and is reflected in the "apparent volume of distribution of the drug". Pharmacokinetic values for the principal cephalosporins3are shown in the table. Department of Bacteriology, Westminster Medical School, London SW1P 2AR
***This letter has been shown whose reply follows.-ED. L.
SYDNEY SELWYN
to
Dr Wise and Dr
Gillett,
SIR,-Firstly, to answer Dr Selwyn’s comment on the lack of detectable cefoxitin in the tissue transudate at 2 z 3h. If all four drugs are compared it is immediately apparent that the tissue levels are closely related to the serum level (and, therefore, serum half-life). The 3 h level of cefoxitin would, therefore, by extrapolation, be 1-2 flg/ml which, as we state, is below the sensitivity of the assay. It is inappropriate to compare cephalexin and CGP 9000, given orally and having a peak serum concentration at 2 h (absorption continuing for some time thereafter), with intravenous cefoxitin. Serum halflives are calculated from the elimination phase. We agree that the free level of an antibiotic in the tissue fluid is of great relevance. We are studying this subject, using a more versatile blister technique to measure both free and bound drug. However, it is of considerable interest that high protein binding does not seem to affect the penetration and concentration of total antibiotic in the tissue. This is an important observation in its own right. Dr Selwyn is incorrect in thinking that "a long serum halflife indicates a relative inability of the drug to leave the bloodstream and enter other body compartments". A well-distributed drug is often more slowly eliminated because these "deeper" compartments act as a depot. Good examples of welldistributed drugs are trimethopriml and griseofulvin,2 with serum half-lives in excess of 9 h. The serum half-life may well (but not necessarily) be of importance in determining an antibiotic’s penetration into tissues. Department of Medical Microbiology, RICHARD WISE Dudley Road Hospital, A. P. GILLETT Birmingham B18 7QH
MUCOCUTANEOUS REACTIONS DURING MYCOPLASMA PNEUMONIÆ INFECTION
SIR,-Dr Lind (March 25, p. 655) found that, among patients with raised cold agglutinin titres, Stevens-Johnson syndrome and non-specific exanthemas were more common in those with positive Mycoplasma pneumoniae serology. 5. Gower, P. E., Dash, C. H. Eur. J. clin. Pharmac. 1977, 12, 221. 1. Fowle, A. S. E. in Trimethoprim/sulphamethoxazole in Bacterial Infections (edited by L. S. Bernstein and A. J. Salter); p. 63. Edinburgh, 1973. 2. Rowland, M., Reigelman, S., Epstein, W. L. J. pharm. Sci. 1968, 57, 984.
1105 We have studied all children referred to Bristol hospitals during an M. pneumonia epidemic lasting 18 months. 9 (20%) out of 44 children with serologically proven M. pneumoniae infection had rashes. 6 children had an erythematous macular rash on the trunk and limbs, and this was usually fading at the time of referral. In 1 of the 6 Stevens-Johnson syndrome developed and there were 3 additional cases of the syndrome. In 3 of the 4 cold agglutinins were sought and all 3 were positive. These findings do not clarify the role of cold agglutinins in the pathogenesis of mycoplasma-associated mucocutaneous reactions. 1 of the children with Stevens-Johnson syndrome, a girl of 11 years, had extensive toxic epidermal necrolysis with typical "scalded skin" erythema, blistering, and epidermal stripping. This rare complication caused significant fluid depletion, but the skin healed within 14 days with no further major problem. The conjunctival ulceration, however, caused healing and fibrosis of such intensity that severe conjunctival shrinkage syndrome, corneal scarring, and visual impairment ensued. 1 case of Stevens-Johnson syndrome was not associated with lower-respiratory-tract infection, which supports Lind’s view that M. pneumonia infection can cause rashes without clinical evidence of respiratory disease.
Cherry et al.2 found no published cases of Stevens-Johnson syndrome associated with M. pneumoniae in girls; 2 of our cases were
females.
Although patients with Stevens-Johnson syndrome usually recover fully we recommend that, in view of the possibility of permanent eye damage, all children with the syndrome should be referred to an ophthalmologist. Royal Devon and
Exeter
Hospital,
Exeter EX2 5DW
Gloucestershire Royal Gloucester GL1 3NN
MEASUREMENT OF LACTOFERRIN IN PANCREATIC JUICE described a protein in pancreatic juice of calcifying pancreatitis;’ this was later identified as lactoferrin,2 a glycoprotein mainly secreted by mammary glands and found in various external secretions.3 Indirect
SIR,-In 1971
men
we
with chronic
immunofluorescence studies revealed lactoferrin in acinar cells of normal pancreatic tissue and in acinar cells of men with chronic calcifying pancreatitis.4 However, the fluorescence was more striking in pathological material. Dr Fedail and colleagues (Jan.. 28, p. 181) have confirmed that lactoferrin secretion is increased in pancreatic juice of patients with chronic
pancreatitis. Since lactoferrin secretion is not increased in pancreatic in acute pancreatitis,s a practical and accurate assay for lactoferrin in pancreatic juice might be a useful diagnostic test. We have compared a double-diffusion technique with a cancer or
radioimmunoassay. The radioimmunoassay was done with pure human lactoferrin (kindly given by Dr P. Masson) labelled with 125by the chloramine-T method. Separation of bound and free lactoferrin was achieved by a double-antibody technique. The sensitivity of the assay was 1 ng/ml. The antilactoferrin prepared in the laboratory was used after a dilution of 1/50 000. ’Immunoplates LC’ (low-concentration) ’Partigen’ lactoferrin and pure standard lactoferrin (titrating 4 mg/dl) were kindly prepared by Dr Baudner (Behring Institute,. Marburg).
1.
PETER G. F. SWIFT
Hospital, DAVID W. STEVENS
Clemente, F., Ribeiro, T., Colomb, E., Figarella, C., Sarles, H. Biochim. biophys. Acta, 1971, 251, 456. 2. Colomb, E., Estevenon, J. P., Figarella, C., Guy, O., Sarles, H. ibid. 1974, 342, 306. 3. Masson, P. La lactoferrine. Paris, 1970. 4. Colomb, E., Pianetta, C., Estevenon, J. P., Guy, O., Figarella, C., Sarles, H. Digestion, 1976, 14, 242. 5. Estevenon, J. P., Sarles, H., Figarella, C. Scand. J. Gastroent. 1975, 10, 327.
SERRATIA MARCESCENS COLONISING THE GUT
SIR,-During an outbreak of hospital cross-infections by multiply resistant Serratia marcescens this organism was occasionally recovered from stools as well as from the site of clinical infection. The spread of Klebsiella pneumoniae from a gastrointestinal-tract reservoir to extraintestinal sites is well documented,’ but S. marcescens does not usually colonise the gut of adults.2 We extended our observation by taking rectal swabs from patients admitted during the outbreak; the swab was streaked on selective media and the plate was inspected caused
after 24 and 48 h incubation at 37°C; the isolates were identified and antibiotic susceptibilities were determined by standard methods. 120 rectal cultures were obtained from 57 patients. The stools of 5 patients were colonised with multiply resistant S. marcescens. 2 patients were colonised or infected at extraintestinal sites at the time the first stool culture was obtained; the other 3 patients later became colonised or infected at extraintestinal sites. As with K. pneumonice, gastrointestinal colonisation by S. marcescens may, by serving as a reservoir for cross-infection, be an important step in the development of hospital-acquired infection. Infectious Disease Section, Department of Medicine, Veterans Administration Hospital, University of Oregon Health Science Center, Portland, Oregon 97207, U.S.A.
STEPHEN R. JONES MERIDITH AMON CATHERINE FALVEY KATHRYN PATRICK
Comparative 1.
Stevens, D, Swift, P. G. F., Johnston, P. G. B., Kearney, P. J., Corner, B. D., Burman, D. Archs Dis. Childh. 1978, 53, 38. 2 Cherry, J. D., Hurwitz, E. S., Welliver, R. C. J. Pediat. 1975, 87, 369. 1. Selden, R., and others Ann. intern. Med. 1971, 74, 6737. 2. Farmer, J. J, and others Lancet, 1976, ii, 455.
measurement of lactoferrin.
Radioimmunoassay (e);
assay
by
radial immunodiffusion
(0).
Abs-
cissa ; pancreatic juice from patients with chronic pancreatitis (1-6) and controls values.
(7-12).
The dotted line represents the limit of normal
