6-aminopenicillanic acid

Sensitisation data (human)

Administrative data

Endpoint:

sensitisation data (humans)

Type of information:

other: case studies

Adequacy of study:

supporting study

Reliability:

other: Reliability scoring in the regulatory sense is not applicable.

Rationale for reliability incl. deficiencies:

other: Reliability scoring in the regulatory sense is not applicable.

Data source

Reference

Reference Type:

publication

Title:

Asthma due to inhaled chemical agents: ampicillin, benzyl penicillin, 6 amino penicillanic acid and related substances.

Author:

Davies RJ, Hendrick DJ & Pepys J

Year:

1974

Bibliographic source:

Clinical Allergy, 4: 227-247

Materials and methods

Type of sensitisation studied:

respiratory

Study type:

other:

Test guideline

Qualifier:

no guideline available

Principles of method if other than guideline:

An investigation into the occurance of asthma in workers employed in the production of antibiotics. Four workers were included in the study, three of which suffered from asthma. The asthmatic patients underwent an inhalation challenge test. Patients were tested with a purified and a commercial grade to include the effects of any impurities.

GLP compliance:

not specified

Test material

Test material information

Constituent 1

Reference substance name:

6-aminopenicillanic acid

EC Number:

208-993-4

EC Name:

6-aminopenicillanic acid

Cas Number:

551-16-6

Molecular formula:

C8H12N2O3S

IUPAC Name:

6-amino-3,3-dimethyl-7-oxo-4-thia-1-azabicyclo[3.2.0]heptane-2-carboxylic acid

Constituent 2

Reference substance name:

Ampicillin

EC Number:

200-709-7

EC Name:

Ampicillin

Cas Number:

69-53-4

IUPAC Name:

6-{[amino(phenyl)acetyl]amino}-3,3-dimethyl-7-oxo-4-thia-1-azabicyclo[3.2.0]heptane-2-carboxylic acid

Details on test material:

- Name of test material : 6-APA or 6-aminopenicillanic acid
- Analytical purity: No data

- Name of test material : Ampicillin
- Analytical purity: No data

Method

Type of population:

occupational

Ethical approval:

not specified

Remarks:

Not reported within study

Subjects:

- Number of subjects exposed: 4
- Sex: All subjects were male

PATIENT 1:
- Age: 39
- Demographic information: A maintenance engineer who had been employed from the age of 33.


PATIENT 2:
- Age: 26
- Demographic information: Employed in the extraction department from the age of 23.


PATIENT 3:
- Age: 44
- Demographic information: Employed in the production of penicillin antiobiotics from the age of 42.


PATIENT 4:
- Age: 39
- Demographic information: Employed in the extraction department from the age of 27.

Clinical history:

- History of allergy or casuistics for study subject or populations:

PATIENT 1
- Symptoms, onset and progress of the disease: Symptoms began 2 years after first exposure. Primarily he developed sneezing and wheezing. The symptoms appeared to be augmented with contact to ampicillin. 15 to 30 minutes after exposure he developed sneezing and wheezing lasting for approximately 1 hour following which he would improve, however he reported waking with a cough and tightness in the chest in the early hours the following morning, with these latter symptoms being the chief complaint. Improvement was noted at weekends, however after a prolonged period without exposure (holiday), the patient found it took 3-4 days before he was entirely free of symptoms.
- Exposure history: The patient worked with ampicillin and related compounds.
- Family history: One daughter with asthma
- Medical history: No previous history of asthma, hay fever or rhinitis.
- Any other allergic or airway disorders: No known drug allergies.
- Smoking history: 10 cigarrettes a day

PATIENT 2
- Symptoms, onset and progress of the disease: After 4 months of exposure he developed an erythematous dry rash on his hands and face which cleared spontaneously but occasionally recurred. After moving to a department with less penicillin dusts developed an attack wheezing within a month occurring several hours after a temporary secondment to the 6-APA prodution line. After 4 weeks, he returned wot work and developed a further attach 8 hours after entering the workplace. He was transferred into a "penicillin free" building and suffered a thrid attack after coming into contact with workers from the extraction departement during a break.
- Exposure history: For the first four months of employment, the subject was based in the extraction department mostly in contact with 6-APA. 18 months after the onset of symptoms the patient was transferred to a sector with less exposure to penicillin dusts and later transferred to a "penicillin free" building but still was exposed through contaminated clothes of other workers.
- Family history: A daughter with eczema and his father was said to have had a "penicillin allergy".
- Medical history : No previous history of asthma, eczma or rhinitis.
- Any other allergic or airway disorders: No previous history of eczma or drug allergies
- Smoking history: 30 cigarettes a day

PATIENT 3
- Symptoms, onset and progress of the disease: 2 years after first exposure to penicillin antibiotics the patient developed conjunctivitis. Symptoms were abated with the use of a protective hood. After a further two months, the patient developed eczema on his hangs and after a further month, rhinitis and asthma. It was noted that wheezing began 6-8 hours after stating work, and resolved within 2 hours of returning home. The symptoms became more severe necessitating 3 days off work. After being transferred to a "penicillin free" building and remained without symptoms for 1 month before developing further attacks of cough and wheeze.
- Exposure history: Initially the patient was exposed to particularly fine dusts of benzyl penicillin and E. coli amidase preaperation, rarely encountering 6-APA and ampicillin. Later he was moved to a "penicillin free" building.
- Family history: One of his two sone had asthman and his daughter, eczema.
- Medical history : The subject had no previous history of asthma or rhinitis
- Any other allergic or airway disorders: No previous history of eczema or drug allergy.
- Smoking history: 10 cigarettes a day

PATIENT 4
- Symptoms, onset and progress of the disease: Four years after commencing work, the subject developed shortness of breath on walking quickly on the flat, and a daily productive cough. Heavy exposure to dusts of any nature made his breathing worse.
- Exposure history: From the age of 27, the subject was exposed to benzyl penicillin and 6-APA dusts.
- Family history: No family history of asthma, eczema or rhinitis
- Medical history : No history of wheezing or sneezing
- Any other allergic or airway disorders: No history of eczema or drug allergies
- Smoking history: 20 cigarettes a day

Controls:

All patients had at least one control day when inhalation testing was carried out with the lactose vehicle alone. The results were recorded as the maximum percent fall from the FEV1 prior to exposure to the lactose. Patients were also tested with benzyl penicilloyl polylysine in the skin test.

Route of administration:

inhalation

Details on study design:

TYPE AND DETAILS OF TEST(S) USED
- Skin prick test: A battery of 23 common inhalant and food allergens as well as solutions of ampicillin 10^-2 M, ampicillin polymer 10^-6 M, benzyl penicilloyl polylysine 10^-6 M, minor determinant mixture 10^-2 M, and E. coli amidase solutions at concentrations of 1 to 10 % w/v.
- Body temperature, eosinophil and neutrophil polymorphonuclear leucocyte counts: Oral body temperature was taken before, 1 hour and 8 hours after each provocation test. Eosinophil and neutrophil polymorphonuclear leucocyte counts were preformed from blood taken before, 8 hours and 24 hours after tests.
- Inhalation challenge tests: Analar lactose was administered as a control and a vehicle. The subjects were exposed to commercial ampicillin and commercial 6-aminopenicillanic acid prepared by E. coli amidase, benzyl penicilloyl polysine and 6-aminopenicillanic impurities. Samples of commercial 6-APA and commercial ampicillin had been further purified by an absorption process to remove high molecular weight protein impurities and referred to as "purified" 6-APA and ampicillin. Ampicillin manufactured from penicillin by a partly chemical method not involving E. coli amidase was used and referred to as ampicillin (chemical).
All patients were inititially challenged with the control (lactose). The lactose was dried by heating overnight at 105 C, placed in two plastic trays and tipped backwards and forwards from one tray to the other 0.3 m below. This was performed in a confined space for 30-60 min. During the test the patient wore a disposable surgical cap, gown and gloves in order to prevent continued exposure after completion of the challenge. Provocation challenge with the test material was carried out in exactly the smae way with a the test material added to the vehicle. 10 mg was initially added to the lactose, with tenfold increases until a measurable reaction occurred. All test materials were white powders and were indistinguishable to the patients. Respiratory function was monitored using a dry spirometre (Vitalograph) to measure FEV1 and FVC and a Wright's metre to measure peak expiratory flow rate. The measurements were made before challenge, every 10 minutes after the challenge for the first hour, and then hourly for the rest of the day, and when necessary during the evening and night.
- Oral challenge test: The patient swallowed two capsules containing in total 500 mg of commercial ampicillin or benzyl penicillin. In each case this was a sample of the same material from the inhalation challenge test.
- Pharmacological agents: The effects on the induced asthmatic reactions of inhaled isoprenaline sulphate 0.16-0.24 mg, sodium cromoglycate 40 mg, and beclomethasone dipropionate were given 30 minutes before challenge testing and also, in patient 1, 3-hourly thereafter for 12 hours. In patient 2, isoprenaline was administered on separate occasions 10 minutes before challenge, 1 hour after allergen challenge and during the actual asthmatic reaction. The results of administering sodium cromoglycate and beclomethasone dipropionate both before and 3-hourly post challenge were recorded.

Results and discussion

Results of examinations:

- Patient 1: No abnormalities were noted on physical examination. Haemoglobin, white cell count and differential were normal. The patient's chest X-ray was also normal. Prick testing was positive with house dust, Dermatophagoides farine and D. pteronyssinus and negative with ampicillin and related substances. The patient was admitted to the hospital on three occasions; October 2-9 1972, December 12-21 1971 and January 8-13 1973. During the inhalation challenge, commercial ampicillin, purified ampicillin and chemical ampicillin all produced strong late asthmatic reactions. Provocation challenge with commercial 6-APA and purified 6-APA produced only mild late falls in the FEV1 of 16 and 23% respectively. No asthmatic reactions were observed with benzyle penicilloyl polysine at 20 mg. No astmatic reaction occured during the oral challenge. When administered with the pharmacological agents, neither 40 mg of sodium cromoglycate nor 200 µg of beclomethasone dipropionate administered 30 minutes before challenge blocked the late astmatic reaction. Administration of 200 µg of beclomethasone dipropionate both before and up to 12 hours (at 3-hourly intervals) after the challenge blocked the late asthma, 40 mg sodium sodium cromoglycate had no definite inhibitory effect.

-Patient 2: No abnormalities were noted on physical examination. Haemoglobin, white cell count and differential were normal. The patient's chest X-ray and urinalysis were also normal. Skin testing was found to be negative to the 23 inhalant and food allergens as well as to ampicillin and related substances. The patient was admitted to hospital on two separate occasions; December 1-23 1971 and January 23-February 3 1973. In the inhalation challenge, commercial ampicillin and chemical ampicillin produced late strong asthmatic reactions, however this was not observed with purified commercial ampicillin (which produced only a mild asthmatic reaction). Commercial 6-APA induced a strong asthmatic reaction, however none was produced by provocation challenge with the same amount in its purified form. The asthmatic reaction induced by 1 g of commercial benzyl penicillin was prolonged, FEV1 reached approximated pre-challenge levels after 4 days. Provocation with purified benzyl penicillin was not performed. 6-APA protein impurities, benzyl penicilloyl polylysine and E. coli amidase extracts gave negative results. Along with asthmatic reactions, the patient also experienced sneezing, rhinitis progressing to a blocked nose, and a mild pruritic erythematous rash over the exposed skin of the face, hands and neck (the latter taking 48 hours to resolve). In the oral challenge, administration of two capsules containing commercial ampicillin gave a late asthmatic reaction similar to that observed in the inhalation test, accompanied with gastrointestinal reactions. One hour post ingestion the patient experienced diarrhoea and colicky abdominal pains taking 10 hours to clear. Inhalation challenges were carried out with either commercial ampicillin or commercial 6-APA (for 30 minutes). 40 mg of sodium cromoglycate administered 30 minutes before the allergen failed to inhibit the asthmatic reaction, though this was blocked by similar pre-challenge inhalation of 200 µg of beclomethasone dipropionate. Inhalation of 0.16 mg of isoprenaline sulphate 10 minutes prior to challenge inhibited the late asthmatic reaction. Administration of the same dose of isoprenaline of a separate occasion, 1 hour after allergen challenge blocked the expected asthmatic reaction. The administration of 0.24 mg of isoprenaline sulphate reversed the late asthmatic reaction.

- Patient 3: No abnormalities were noted on physical examination. Haemoglobin, white cell count and differential were normal. The patient's chest X-ray was normal. Skin testing was negative to the 23 inhalant and food allergens as well as to ampicillin and related substance. The patient was admitted to hospital from February 1-18 1973. In the inhalation test, both ampicillin and benzyl penicillin, in commercial and purified forms produced late asthmatic reactions. Chemical ampicillin was not tested. Neither commercial nor purified 6-APA produced asthma on inhalation challenge. Rhinitis occurred at the same time as the late asthmatic reactions and a mild pruritic erythematous rash developed over the exposed area of the face and neck taking up to 48 hours to resolve. In the oral challenge, two capsules containing 500 mg of benzyl penicillin produced a late asthmatic reaction begining only 30 minutes after ingestion and lasting 24 hours. This was accompanied by extensive urticaria but not by diarrhoea.

- Patient 4: No abnormalities were noted on physical examination. Haemoglobin, white cell count and differential were normal. The patient's chest X-ray was normal. Skin testing was negative to the 23 inhalant and food allergens as well as to ampicillin and related substances. The patient was admitted to hospital from November 5-10 1972. In patient 4 there was no evidence of significant reversible airways disease. Inhalation challenge with 10 mg of commercial ampicillin and 10 g of commercial benzyl penicillin for 30 minutes was followed by falls in FEV1 of 8 and 11% only comparable with the 10% on control testing with the lactose vehicle.

Any other information on results incl. tables

Full tabulated results for all the patients are included in the attached appendix, Appendix 1 Tabulated Data for Davies RJ et al 1974.pdf

Generalised Results

 

Controls: During some of the control days, the FEV₁ fell by over 15%. In patient 1, during the first control day only 3 days after factory exposure to ampicillin, the patient’s FEV₁ fell by 16%. The maximum fall occurred during the night, 16 hour post exposure to the control lactose vehicle. However after 2 months away from the factory, his FEV₁ following a further control challenge fell at its maximum 16 hours later by only 2%. Similarly, following a positive asthmatic reaction to ampicillin, the FEV₁ on the following day fell substantially below that no previous control days. It was only on the second post challenge day that the FEV₁ approximated control levels.

 

Pre-challenge FEV₁: The variation above and below the mean did not exceed 13 %.

 

Provocation challenge tests: The induced asthmatic reactions were late in type. Airway obstruction developed slowly, beginning usually 1 hour or more after provocation challenge and was prolonged with maximum falls in FEV₁ occurring between 3 and 16 hours later.

 

FEV₁/FVC ratio: During the strong late asthmatic reactions the FEV₁/FVC ratio fell in patient 1 from 74 to 61% December 19^th 1971, in patient 2 from 72 to 55 % January 31^st 1973 and in patient 3 from 52 to 35% February 5^th 1973.

 

Body temperature: Oral temperature did not rise above normal in any of the provocation tests in all of the subjects.

 

Eosinophil and neutrophil polymorphonuclear leucocyte count: In Patients 1, 2 and 3 blood eosinophilia occurred within 24 hours of the first positive provocation challenge. The counts tended to increase in positive asthmatic reactions occurred on subsequent days, and decrease in the absence of the reactions. No blood eosinophilia occurred in patient 4. None of the patients developed an increase above normal in the neutrophil polymorphonuclear leucocytes.

Applicant's summary and conclusion

Conclusions:

Three out of four workers from an antibiotic-producing factory developed late asthma and eosinophilia on inhalation challenge testing with ampicillin and related substances. The chemicals to which each worker reacted varied, suggesting differences in the clinically important allergens for each individual.
Oral dosing with ampicillin in one patient, and benzyl penicillin in another in therapeutic doses led to the development of late asthma accompanied by skin and gastro-intestinal disturbances.
The inhalation of 200 µg of beclomethasone dipropionate before in one case, and before and 3-hourly after provocation challenge in the other, blocked the asthmatic reaction. 40 mg sodium cromoglycate similarly given failed to inhibit the reactions. In one patient, 0.16-0.24 mg of isoprenaline sulphate inhaled 10 minutes before or 60 minutes after inhalation testing prevented the development of late asthma, and when given during the asthmatic reaction led to its rapid reversal.
The three patients differed in respect to the substances which produced late asthmatic reactions. Patient 1 developed a strong late asthmatic reaction to “purified” ampicillin as well as ampicillin (chemical). On the other hand the reactions developing with commercial or “purified” 6-APA were mild. The absence of any difference in the asthmatic reactions to commercial or “purified” products or to ampicillin produced be a different method is evidence against impurities being important in the aetiology of these late asthmatic attacks. The very slight reactions to 6-APA suggest that the important allergen for this patient seems to be in relation to the extra side chain linked to the 6-APA nucleus. Patient 2 developed late asthmatic reactions on exposure to commercial ampicillin and ampicillin (chemical) but only very slight reactions to “purified” ampicillin. Although a late asthmatic reaction occurred with commercial 6-APA, none followed exposure to purified 6-APA suggesting that the impurity is the allergen. The patient suffered a reaction to commercial benzyl penicillin, 6-APA is produced commercially from benzyl penicillin with E. coli amidase enzyme. Protein impurities including remnants of E. coli enzymes were considered possible allergens, however in this patient chemical benzyl penicillin and ampicillin induced late asthma. 6-APA protein impurities and E. coli amidase extracts were tested in small amounts which proved negative, however the small volume of test material may have been insufficient to induce a reaction. Patient 3 developed late asthmatic reactions when exposed to ampicillin and benzyl penicillin, whether commercial or purified, but no reaction occurred with commercial or purified 6-APA. This suggests that the important allergens for this patient are related to the structure of ampicillin and benzyl penicillin rather than impurities.