Glutaral

Endpoint summary

Description of key information

Several data on the sensitizing potential of glutaraldehyde are available, tested with an open cutaneous test, LLNA, Guinea Pig Maximisation Test, Buehler test, and the mouse ear swelling test. Besides a Buehler test and one guinea pig maximisation test, all studies indicate that glutaraldehyde is sensitising to skin. In addition, in human studies sensitising effects on the skin and respiratory tract were observed.

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Reference

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1975

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

The study was conducted prior to the implementation of the actual guidelines and GLP was not compulsory at the time the study was performed. However, the study was based on an acknowledged standardized test method and was conducted according to an acceptable BASF test; basic data were given.

Principles of method if other than guideline:

BASF test:
The aim of the present test was to look for the sensitizing potential of glutaraldehyde using the open epicutaneous test with guinea pig, which is an acknowledged standardized test method.
For induction, the unchanged test substance was applied to the shaved left flank (area 25 cm2) of 10 Pirbright White Guinea pigs. Application was performed once daily, for 5 consecutive days, over 2 weeks (total of 10 applications). The shaved right flank remained untreated.
Following an 11-day recovery period, both flanks were shaved once again and the unchanged test substance was applied, this time to the right flank of each treated animal (single application). Three control animals, which had not been previously subjected to induction, were treated similarly. Skin reactions were recorded after 12 hours.

GLP compliance:

no

Remarks:

; GLP was not compulsory at the time the study was performed.

Type of study:

open epicutaneous test

Justification for non-LLNA method:

The study was performed in 1975, before the LLNA OECD TG 429 was in place (adopted in 2002).

Specific details on test material used for the study:

- Name of test material (as cited in study report): Glutaraldehyde 25% aq. solution
- Physical state: liquid
- Analytical purity: 25.0% glutaraldehyde

Species:

guinea pig

Strain:

other: Pirbright White guinea pigs

Sex:

female

Route:

epicutaneous, open

Vehicle:

unchanged (no vehicle)

Concentration / amount:

The application area was a skin area of 25 cm2

Route:

epicutaneous, open

Vehicle:

unchanged (no vehicle)

Concentration / amount:

The application area was a skin area of 25 cm2

No. of animals per dose:

Treated females: 10
Control females: 3

Details on study design:

INDUCTION EXPOSURE
- No. of exposures: 10 applications
- Exposure period: 2 weeks
- Test groups: one group comprising 10 animals
- Site: left flank, clipped 25 cm2 area within the fore region; the shaved right flank remained untreated

CHALLENGE EXPOSURE
- No. of exposures: single application
- Day(s) of challenge: challenge was conducted after a recovery period of 11-day following induction
- Exposure period: 12 hours
- Test groups: one induced group comprising 10 animals
- Control group: Three control animals, which had not been previously subjected to induction, were challenged.
- Site: both flanks were shaved again and the unchanged test substance was applied, this time to the right flank of each treated animal
- Evaluation (hr after challenge): after 12 hours following challenge

Reading:

1st reading

Hours after challenge:

12

Group:

test chemical

Dose level:

single application of glutaraldehyde 25% on a skin area of 25 cm2

No. with + reactions:

10

Total no. in group:

10

Clinical observations:

All animals showed slight spot-like redness of the skin and brownish residues of the test substance.

Remarks on result:

positive indication of skin sensitisation

Reading:

1st reading

Hours after challenge:

12

Group:

negative control

Dose level:

single application of glutaraldehyde 25% on a skin area of 25 cm2

No. with + reactions:

0

Total no. in group:

3

Clinical observations:

None of the control animal showed skin reaction. Brownish test substance residues on the skin were seen.

Remarks on result:

no indication of skin sensitisation

Repeated induction treatment with the undiluted test substance caused necrotic skin lesions with thick bloody
crusts in all animals. Upon challenge treatment of the opposite flank with the undiluted test substance performed
after an 11-day period without treatment, all animals exhibited slight erythema. No signs of irritation were seen
in the three untreated controls.

Conclusions:

Glutaraldehyde 25% was sensitizing to guinea pig when tested in the open epicutaneous test.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (sensitising)

Additional information:

Open epicutaneous test

An Open Epicutaneous Test with guinea pigs was conducted with a 25% aqueous solution of glutaraldehyde containing about 8% methanol and 67% water (BASF, 1975). For induction, the neat test material was applied to the shaved left flank of 10 Pirbright White Guinea pigs; the shaved right flank remained untreated. Application was performed once daily, for five consecutive days, over 2 weeks (total of 10 applications). The treatment resulted in thick bloody scabs. Challenge was conducted after a latency period of 11 days and consisted of a single application of the undiluted test material to the right flank of each treated or control animal. Skin examination after 12 hours following challenge revealed slight spot-like redness of the skin for all treated animals whereas the controls showed no skin reaction. These results are indicative of a skin sensitizing potential for glutaraldehyde 25%, and a similar sensitizing potential has to be expected for glutaraldehyde 50%.

LLNA

The sensitizing effect of glutaraldehyde was also assessed in a LLNA (Ulrich, 2001). The study mainly focused on cytokine response patterns following sensitization to address a possible application of cytokines as markers for secondary contact allergy. The authors reported that glutaraldehyde treatment resulted in a significant increase in ear weight in comparison to the induction control groups, coupled with significant increase in lymph node weight. Furthermore upon challenge, glutaraldehyde treatment resulted in a marked up-regulation of the IL-4 gene expression and an increased synthesis of interleukin IL-4. CD4+T cells were identified as the main cell type responsible for IL-4 release. In contrast, interferon-Beta (IFNß) was neither up- nor down regulated. The comparison of the cytokine responses to skin and respiratory sensitizers revealed no evidence for specific response patterns. In conclusion, (1) glutaraldehyde was assessed in this assay as a moderately active contact allergen, (2) skin sensitizers are able to elicit cytokine response patterns and particularly T helper 2 cytokines (e.g. IL-4) measurement after challenge may by considered as a stable marker, and (3) no particular preference of cytokine response patterns for either dermal or respiratory sensitizers was evident.

The Dow Chemical Company (1994) exposed four mice with dose levels of 0, 5, 10, 25, and 50%, respectively. Each of the test concentrations produced substantial levels of lymph node cell proliferative activity. Increasing concentrations of the test material elicited stimulation indices of 15.5, 23.4, 38.7, and 34.9. The data confirms that the test material produces skin sensitization potential. An EC3 was not calculated.

Hilton and co-workers (1998) investigated the sensitizing potential of glutaraldehyde and formaldehyde in a comparative manner using the LLNA; the possible influence of the used vehicle was also considered. In both vehicles, acetone and DMF, glutaraldehyde induced substantially more vigorous responses in the local lymph node assay (EC3 values of 0.006mol/L in acetone and 0.002mol/L in DMF) than did formaldehyde (EC3 values of 0.18mol/L in acetone and 0.11mol/L in DMF). These results demonstrate that glutaraldehyde has a considerably greater potential to induce skin sensitization than does formaldehyde; the data are consistent with what is known of the ability of these chemicals to cause allergic contact dermatitis in human.

In a study performed by Azadi (2004) several immunological responses following glutaraldehyde exposure were investigated. Among the investigations was a LLNA test in which 5 female Balb/c and CBA mice were topically exposed to the vehicle or 0.1, 0.75, 2.5 % glutaraldehyde, or positive control (HCA 30%) on the dorsal surface of each ear (25 µl per ear) for three consecutive days. On day 6, mice were injected intravenously via the lateral tail vein with 20 µCi 3H-thymidine. Five hours after 3H-thymidine injection, animals were euthanized via CO2 inhalation, and the left and right cervical draining lymph nodes (DLNs) located at the bifurcation of the jugular vein were excised and pooled for each animal. In CBA mice, exposure to all concentrations tested resulted in stimulation indices of >3, ranging from 4.9 to 31.5, with an EC3 value of 0.072%. In Balb/c mice exposure to to glutaraldehyde resulted in stimulation indices ranging from 3.5 to 25.5, with an EC3 value of 0.089%. The positive control group (30% HCA) resulted in a SI of 12.3 in CBA and 8.7 in BALB/c mice. No signs of systemic toxicity were observed at concentrations up to 2.5% glutaraldehyde.

In a LLNA study (Van Triel, 2011) 6 male BALB/c mice per dose received 0.25% or 2.5% GA dissolved in acetone olive oil (4:1) on the skin of the ears (25 µL) on three consecutive days. At necropsy, mandibular and auricular LNs were excised. Subsequently, draining LNs were used for determination of proliferation by labeleling ex vivo of DNA with [3H]-thymidine and determine cytokine production by flow cytometry. Mean [3H]-thymidine incorporation was calculated per experimental group. Stimulation indices (SI) were then calculated by dividing the mean [3H]-thymidine incorporation per experimental group by the mean [3H]-thymidine incorporation of the (vehicle-treated) control group. Chemicals that are able to elicit a stimulation index of 3 or more were considered skin sensitizers in the dermal LLNA. A dose-dependent increase in cellular proliferation was found in the auricular lymph nodes, with stimulation indices (SI) of 14 and 33 in response to 0.25% and 2.5% GA, respectively. A Th2-dominant cytokine profile was observed already at the low dermal concentration and the IL-4/IFN-gamma ratio shifted even more towards a Th2-like cytokine expression profile with increasing dose.

In another study summary from Proctor & Gamble (1996) an examination was performed of proliferation in responder lymph node cells treated for use in a blastogenesis assay using the local lymph node assay protocol and stimulator dendritic cell number and percent. For the LLNA assay 5 female Balb/c mice per dose were induced with 0.1, 0.5, and 1 % of 12.5 µL of test material applied one time a day for four days to the dorsal and ventral sides of both ears. Four days after the last induction treatment, the responder cell mice were injected, via the tail vein, with 20 µCi 3H-thymidine in 0.25 mL sterile PBS. At approximately 5 hours after injection of the 3H-thyrnidine, the mice, were sacrificed by CO2 inhalation. The two auricular lymph nodes were excised and pooled for each individual animal in 4 mL PBS. Mean DPM scores and stimulation Indices (S.I.) were, 705.6 (0.8), 1226.6 (1.5) and 2073.0 (2.6) for 0.1, 0.5 and 1% glutaraldehyde respectively. For treatment of 8 mice as donors of stimulator dendritic cells, each mouse was clipped on the dorsal side form the nuchal area to the rump. Mice were treated with 100 µL of the test material on the clipped dorsal area, and 12.5 µL of the test material on each side of both ears for a total of 25 µL per ear. At approximately 24 hours post-treatment, the mice were sacrificed by CO2 inhalation. The auricular, branchial, axillary and inguinal lymph nodes were excised from each mouse and was pooled for each group. Mean cell number at 1% glutaraldehyde was 2.60E-4 / node and the percentage dendritic cells at 1 % glutaraldehyde was 25.9% . The mean cell number for the naive (control) was 1.81E-4/ node and the percentage dendritic cells was 28.7%.

In a study from Takeyoshi (2005), a new procedure was demonstrated for classifying chemicals, among which glutaraldehyde, according to their sensitization potency classes by non-radioisotopic LLNA. Therefore, 25-μl volume of 2 or 10% glutaraldehyde was applied to the dorsum of both ears of female CBA/JN mice daily for three consecutive days. A single intraperitoneal injection (5 mg per mouse per injection) of (5-bromo-2-deoxyuridine) BrdU was given on day 4. The stimulation index was 15 for both concentrations. The authors classified glutaraldehyde as moderate sensitizer.

In a study performed by Yamashita (2015) 5 female CBA:J mice received 1.56% in DMF on the skin of the right ear on day 1, 2, 3 and on both ears on day 10. For each control group, test chemicals were applied to the dorsum of the left ear on day 10. Local lymph nodes were excidses and weighed on day 12. Body weights were recorded on day 1 and 12. Systemic toxicity was observed through the experimental period. - When 1.56% glutaraldehyde were tested, a clear elicitation response was observed. Glutaraldehyde was identified as strong skin sensitizer (DILL value 1.09).

In a study from Hou (2015) that was conducted according to OECD Guideline 442B (Skin Sensitization: Local Lymph Node Assay: BrdU-ELISA) 4 male BALB/c mice per dose received 1 % GA dissolved in acetone olive oil (4:1) on the skin of the ears (25 µL) on three consecutive days. On day 6, a pair of auricular lymph nodes from each mouse was excised. A positive response was defined as the mean SI of the test group of ≥1.6. In this study a mean SI of 2.78 was determined. Therefore, GA was classified as sensitizer.

GPMT

In a study Guinea pig maximisation test by Ballantyne (1997), 10 male and 10 female Dunkin-Hartley guinea pigs were exposed to unbuffered 2.2% glutaraldehyde and to buffered 2.2% glutaraldehyde. Each animal received 6 injections of 0.1 ml each in clipped shoulder region. These injections consisted of 2 injections each of an aqueous 50% suspension of Freund´s Complete Adjuvant, without test substance; 2 injections each of 0.1 ml of 5% test substance in propylene glycol; 2 injections of an aqueous 50% suspension of Freund´s Complete Adjuvant with test substance. The epicutaneous induction was conducted after 1 week following the intradermal induction. The undiluted test substance (2.2% glutaraldehyde) was applied to the clipped shoulder under occlusive conditions. Challenge was carried out 2 weeks later with a 10% dilution. Unbuffered glutaraldehyde 2.2% was sensitising in the GPMT. Alkaline buffered glutaraldehyde 2.2% was sensitising in the GPMT, however, the sensitising potential was lower than for the acidic unbuffered test material.

In a summary report from the EPA (1992), a guinea pig maximisation test was described to screen the test substance for its skin sensitisation potential. For this test, 10 animals were exposed intradermal and epicutaneous. Intradermal induction took place once with 0.1mL of 3% solution of glutaraldehyde in saline. Topical induction took place for 48 hours (occluded) with a 2x4 cm patch with 25% glutaraldehyde in saline solution. The epicutaneous challenge took place for 24 hours under occlusive conditions with a 2x2 cm patch containing 10% glutaraldehyde in saline. When evaluated under the condition of the Magnussen-Kligman sensitization test method, the test substance elicited no positive responses indicative of sensitization.

Buehler test

Nutrition International Inc. (1982) tested glutaraldehyde in a Buehler test using 12 male Hartley guinea pigs. Each animal received a total of 9 applications (occlusive) on alternate days for induction. Each application lasted for 6 hours, thereafter the patches were removed and the application sites were gently cleaned. On the basis of the pre-test a test concentration of 10% was first selected for induction. However, following the first application, unexpected erythema was seen in 4 of the 12 test animals, therefore the test concentration was reduced to 3% for the further applications. As the concentration of 3% also produced slight erythema after the second and the third application, the concentration was further reduced to 1% for all subsequent applications. Challenge was conducted after 14 days following the last application of the induction series with 1%. The test material applied as a 1% solution to guinea pigs did not produce skin sensitization.

Mouse ear swelling test (MEST)

In a sensitization assay by Gad et al. (1986) the mouse ear swelling test (MEST) design was used to evaluate 72 materials (among which glutaraldehyde). Female CF-1 mice were shaved and tape stripped at the start of the study. Prior to the first application of 100 µL of a 1 % solution of the test substance (in 70 % EtOH aq.), mice received injections totaling 0.05 mL Freund's Complete Adjuvant (FCA) into the stomach induction site. After application of the test substance, the application site was allowed to dry. Tape stripping and topical applications were repeated for three additional days. Seven days after the last topical application to the stomach, 20 µL of the test solution was applied to the left ear of the mice and another 20 µL to the right ear. At 24 and 48 hour after this challenge, animals were lightly anesthetized with ether, and the thickness of both ears measured. Results were expressed as percentage of animals sensitized to the test substance and percentage ear swelling compared to the controls. Of the tested animals 67 % were sensitized to the test substance with a mean of 125 % ear swelling compared to the controls.

In a MEST by Descotes (1988) mice received an epicutaneous application of the test substance (1%) on both sides of the right ear on day 0 and 2, as well as a scapular s.c. injection of complete Freund´s adjuvant on day 2. On day 9, the thickness of the ear of the induced animals was measured prior and 24 hours after epicutaneous application of the test substance on both sides of the ear. For glutaraldehyde, ear thickness prior to challenge was 21.56 mm and after 24 hours following challenge the measured value was 26.69 mmm indicating a significant ear swelling (123.8%). Glutaraldehyde was therefore positive in the MEST.

Dunn et al. (1990) investigated the ability of a MEST, previously developed and validated by Gad et al. (1986), to predict contact sensitization induced by weak to moderate contact sensitizers. A series of 26 chemicals was tested including glutaraldehyde in two laboratories. For induction, 100 µL of test material (1%) was applied on the abdominal skin of each mouse and allowed to dry. On 3 consecutive days following the first induction adhesive tape was used to remove stratum corneum and 100µL test chemical was applied topically. On day 9 after induction, the thickness of the ear was measured to assure that the baseline thickness of the ear was within the normal range of ca. 0.19 to 0.22 mm. On day 10, challenge was performed by applying 10 µL test substance (10%) on the dorsal side and 10 µL on the ventral side of the left ear of each mouse.. The right ear was "challenged" with vehicle (ethanol 70%). Each ear was measured for thickness after 24 hours following challenge. As for both laboratories the percentage of animals reacting positively after challenge exceeded 20% (11 of 14 animals and 3 of 14 animals for both laboratories, respectively).

The purpose of the study by Cornacoff (1988) was to evaluate the MEST and to compare MEST with an isotopic modification. Therefore, a radio isotopic incorporation assay utilizing [125I]iododeoxyuridine was compared to the standard mouse ear swelling test (MEST) for a series of sensitisers including glutaraldehyde. The MEST test was conducted according to the method described by Gad et al. (1986). Balb/c mice received two 25 µL- intracutaneous injections of Freund's complete adjuvant within the abdominal region. A volume of 25 µL test substance (10%) was applied onto a filter disc which was attached to an Elastoplast tape strip. The tape strip was then wrapped around the abdomen of the test animal and was secured in place. This first application was followed by 3 further applications on the 3 consecutive days. One day prior to challenge, each animal received an i.p. injection of 1 mg/kg bw of 5-fluorodeoxyuridine (FUdR) followed one hour later by an i.p. injection of 1 µCi of [125I]-iododeoxyuridine (IUdR; 2200 Ci/mmol). Challenge consisted of an application of 25 µL of the test substance (2%) on the left ear; the right ear served for control and received an application of the vehicle. The mice were sacrificed after 24 to 48 hours and the ears were evaluated for thickness and for the infiltration of [125I]-IUdR-labeled cells. Two experiments were conducted with 12 mice each. For glutaraldehyde, no reactivity was detected by the radioisotopic assay, but the MEST resulted in a minimal but significant positive result.

In the same publication by Azadi et al. (2004) the mouse ear swelling test (MEST) was performed. Therefore BALB/c or B6C3F1 mice were exposed topically (induction) on the shaved lumbar area with the vehicle substance, increasing concentrations of glutaraldehyde (50 µL of 0.1, 0.75, 2.5 % solutions), or the positive control substance for 3 days. On day 8, before chemical challenge, the thickness of the right ear pinna of each mouse was measured, mice then received a single challenge exposure of 25 µL vehicle, 2.5% glutaraldehyde, or 0.15% 2,4-dinitrofluorobenzene (DNFB) on the dorsal surface of the right ear pinna. Ear measurements were taken 1/2, 24, and 48 h following challenge. An immediate response was observed in animals induced and challenged with 2.5% glutaraldehyde whereas animals induced with 0.1% or 0.75% and challenged with 2.5% exhibited a delayed response 48 h post challenge. Results suggested an immediate (Th2) response in the 2.5% exposed group and a delayed (Th1) response in the 0.1% and 0.75% exposed groups. Furthermore, total serum IgE was significantly elevated (P < 0.01) only in the group of animals exposed to 2.5% glutaraldehyde compared to the vehicle control, further supporting a Th2 response in this group. In addition to the MEST assay, phenotypic analysis was performed to quantify the percentage of B220+- and IgE+B220+-expressing cells in the draining lymph nodes. Mice were exposed to the vehicle N,N-dimethylformamide (DMF) or increasing concentrations of glutaraldehyde topically on the dorsal surface of each ear (25 µL/ear of 0.1, 0.75, 2.5 % solutions) for four consecutive days. Animals were allowed to rest for 6 days after the final exposure and then euthanized on day 10 by CO2 inhalation. Lymph node cell phenotypes were analyzed using flow cytometry. A statistically significant increase (P < 0.01) in absolute and percentage of B220+ cells was seen for all exposure groups. A concentration-dependent increase in the absolute number

( P < 0.01) and percentage ( P < 0.05) of IgE+ B220+ expressing cells was observed following glutaraldehyde exposure reaching statistical significance at 0.75% (P < 0.05) and 2.5% (P < 0.01) exposure levels. IgE levels were elevated with increasing concentrations of exposure reaching significance in animals exposed to 2.5% glutaraldehyde. Finally cytokine mRNA in the draining lymph nodes was analyzed to further evaluate the effect of concentration of exposure on Th1/Th2 balance. Cytokine mRNA levels analyzed included IL-4, -10, -12, and IFN-y. A significant increase in IL-4 mRNA in the draining lymph nodes was observed only in the 2.5% exposed group. These results indicate that the development of an immediate vs. a delayed hypersensitivity response following dermal exposure to glutaraldehyde is at least in part mediated by the exposure concentration.

Human data on skin sensitization

Several human population studies were identified regarding skin sensitization effects after exposure to the test substance.

Ballantyne et al (1984) performed patch tests (induction and challenge) on 109 volunteers with glutaraldehyde at 0.1, 0.2 and 0.5 % in aqueous solution. Glutaraldehyde at concentrations of 0.1% and 0.2% produced no evidence of a sensitization reaction, but at 0.5% there was a definite reaction to the challenge patch in one of the 109 subjects. According to the authors, this indicated that 0.5% glutaraldehyde is around the threshold concentration for induction of dermal sensitization to the test material. A study performed by Seneja et al (2008), in which a total of 1434 subjects underwent patch testing, showed that the prevalence of glutaraldehyde allergy was 15 times greater in health care workers than in non-health care workers. Similarly, in a 5-year study in which 468 subjects were patch tested to glutaraldehyde, Shafer et al (2000) found that health-care workers were more than 8 times more likely to be allergic to glutaraldehyde than their non-health-care working peers. Géraut et al (2007) performed a multicentre study on 1919 patients in order to evaluate sensitization due to glutaraldehyde in a general population tested for contact dermatitis. The authors of the study reported a positive reaction against glutaraldehyde tested in 1% vaseline in 4.1% of cases (i.e. 79 patients). Reifenrath et al (1985) performed an 8-week irritancy test by applying a 10% aqueous solution of glutaraldehyde to the ankle and heel area of 12 volunteers. Irritation and one case of sensitization resulted from glutaraldehyde application to areas of thin stratum corneum (anterior ankle) but not from applications to thick stratum corneum (posterior ankle). Jachuk et al (1989) used a questionnaire to determine the symptoms associated with glutaraldehyde in 9 medical and nursing staff members working in a endoscopy unit. Eight members of the staff, who had been affected by the vapour, also underwent clinical assessment. Manifestations of glutaraldehyde induced symptoms included watering of eyes, rhinitis, breathlessness, dermatitis, nausea, wheezing, and headache. Skin patch tests were not performed. The exposure measurements revealed an exposure of 0.05 - 0.12 ppm. In addition to the above, a review by Takigawa et al (2006), a review by the Dutch Expert committee (2005), and a review by Maier et al (2009) described increased risk of skin sensitization effects after glutaraldehyde exposure.

In contrast to the above, the Union Carbide Corporation (1966) performed a repeated insult patch test on ambulatory volunteers, and elderly people in a nursing home with 5. 2 and 1 % glutaraldehyde solution. There was according to the authors no evidence of sensitizing action. In an epidemiological study from the Union Carbide Corporation (1993, later published by Teta et al 1995) the incidence of skin sensitization, respiratory sensitization and allergic blepharoconjunctivi was examined in 218 men assigned to glutaraldehyde production or drumming from its start-up in 1959 through 1992, at a chemical plant in West Virginia. Plant medical records, work restrictions, and clinic visits for accidental exposures were reviewed. There were no indications of glutaraldehyde-induced skin or respiratory sensitization, or allergic blepharoconjunctivitis in workers exposed at or below 0.2 ppm of glutaraldehyde. Juhlin et al (1986) studied 160 patients in a 6-week sweat inhibition test in which a 10% aqueous solution was applied to the feet (palms) of patients with hyperhydrosis. They were routinely tested for contact dermatitis and were patch-tested with 1% glutaraldehyde. Three applications a week of a 10% solution was sufficient to keep the feet free from excessive sweating. The treatment caused no irritation in the patients. On the palms prolonged treatment was discontinued due to visible discoloration. The patch tests in the 160 patients showed no allergic reaction to glutaraldehyde. In a study by Weaver et al (1977), the potential hazard of adding a low level (550 ppm) of glutaraldehyde to an experimental liquid fabric softener was determined. The subjects were exposed (1) to the experimental formulation, (2) to the formulation with 10 times the normal level of glutaraldehyde, and (3) to fabrics laundered with the normal use level or (4) with 10 times the normal use level of these experimental formulations. No sensitizing effects were observed in any of the experiments.

The following clinical case reports were identified concerning skin sensitization after glutaraldehyde exposure.

Fowler (1998) described the case of a female hospital maintenance employee who developed an airborne contact dermatitis when cleaning respiratory therapy equipment. Patch-testing revealed that the woman was allergic to glutaraldehyde contained in the commercial germicidal product named Cidex (R). Sandersen (1968), described the case of 2 hospital sisters, exposed to gluteraldehyde, suffering from contact dermatitis. The subjects were patch tested with glutaraldehyde (1% aq) and formalin (2%). Both subjects tested positive to glutaraldehyde, and negative to Formalin. Kiec-Swierczynska et al (2001) described the case of 2 hairdressers suffering from dermatitis. The 2 hairdressers showed positive patch test reactions to glutaraldehyde (0.2 % in pet.). Di Prima et al (1988) described the case of a 51 year old male with glutaraldehyde induced contact dermatitis. Subject worked as a cleaner in a radiovascular department and he had developed dermatitis 4 month after Cidex, containing formaldehyde and the test substance, had been introduced for disinfection of surgical instruments. Patch tests with Cidex, Cidex solution (10%), and glutaraldehyde (1 and 0.5%) were positive. Jordan et al (1972) described 5 cases concerning contact dermatitis after glutaraldehyde exposure. Three were dental assistants with occupational exposure and, two were exposed by using a substance for dermatological disorders. Patch tests were performed and all patients reacted to 0.25 and 1% glutaraldehyde in water and to chrome-glutaraldehyde tanned leather. Patch tests to chromate (0.25% in water) and formaldehyde (2.5% in water) were negative. Exposure-tests with leather treated with only chromate or glutaraldehyde were negative for chromate and positive for glutaraldehyde. Fisher (1981 and 1990) also described a total of 4 cases of contact dermatitis after occupational exposure to glutaraldehyde during the handling of x-ray film or during disinfecting of endoscope material with Sporicidin, a glutaraldehyde-phenol formulation. Subjects tested positive when patch tested with a 1% aqueous glutaraldehyde solution and/or patch tested with dilutions Sporicidin. Subjects tested negative to formaldehyde.

Short description of key information:

Several data on the sensitizing potential of glutaraldehyde are available, tested with an open cutaneous test, LLNA, Guinea Pig Maximisation Test, Buehler test, and the mouse ear swelling test. Besides a Buehler test and one guinea pig maximisation test, all studies indicate that glutaraldehyde is sensitising to skin. In addition, in human studies sensitising effects on the skin and respiratory tract were observed.

Respiratory sensitisation

Link to relevant study records

Reference

Endpoint:

respiratory sensitisation

Remarks:

other: Assessment based on Human data

Type of information:

other: not applicable

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Acceptable review

Principles of method if other than guideline:

Not applicable

GLP compliance:

no

Remarks:

; not applicable

Species:

human

Strain:

other: not applicable

Sex:

male/female

Route of induction exposure:

other: not applicable

Route of challenge exposure:

other: not applicable

Vehicle:

other: not applicable

Concentration:

Not applicable

No. of animals per dose:

Not applicable

Results:

According to the review, glutaraldehyde can cause asthmatic symptoms, such as wheezing, coughing, chest tightness, breathing difficulties and non-specific hyperresponsiveness. The asthmatic symptoms were considered to be indicative of a respiratory sensitizing potential for glutaraldehyde. However, at the time the review was published, human and animal data on respiratory tract sensitisation still were not conclusive. In fact, it was suggested that the irritating properties of glutaraldehyde may have resulted in work-aggravated asthma in susceptible individuals, meaning that pre-existing or concurrent asthma will be aggravated by irritants in the workplace. Nevertheless, based on immunological tests in humans and the few animal studies carried out, and due to the fact that glutaraldehyde may cause allergic skin sensitisation, the committee concluded that glutaraldehyde should be considered as a respiratory sensitizer. However, most probably, it should be considered as a respiratory allergen of low potency in view of the large numbers of occupationally exposed persons in relation to the limited number of reported patients. The immunological mechanism of action still remains poorly understood.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (sensitising)

Additional information:

Referring to the respiratory tract sensitizing potential of glutaraldehyde, it was reported that glutaraldehyde can cause asthmatic symptoms, such as wheezing, coughing, chest tightness, breathing difficulties and non-specific hyperresponsiveness. The asthmatic symptoms were considered to be indicative of a respiratory sensitizing potential for glutaraldehyde. However, at the time the review was published, human and animal data on respiratory tract sensitisation still were not conclusive. In fact, it was suggested that the irritating properties of glutaraldehyde may have resulted in work-aggravated asthma in susceptible individuals, meaning that pre-existing or concurrent asthma will be aggravated by irritants in the workplace. Nevertheless, based on immunological tests in humans and the few animal studies carried out, and due to the fact that glutaraldehyde may cause allergic skin sensitisation, the committee concluded that glutaraldehyde should be considered as a respiratory sensitizer. However, most probably, it should be considered as a respiratory allergen of low potency in view of the large numbers of occupationally exposed persons in relation to the limited number of reported patients. The immunological mechanism of action still remains poorly understood.

In a respiratory LLNA study from van Triel (2011), 6 male BALB/c mice per dose were exposed by inhalation to 6 or 18 ppm GA for 1.5 h/day, both generated as a vapor and as an aerosol on three consecutive days. Three days after the last exposure the animals were necropsied and the respiratory tract was collected for histopathological examination. Mandibular and auricular lymph nodes (LNs) were collected, LN cells were harvested, and DNA was labelled with [3H]-thymidine to determine proliferation. In addition, production of several cytokines by the cultured LN cells was measured using a flowcytometer. Immediately after the first exposure, all animals exposed to GA by inhalation were lethargic and exhibited a decreased breathing frequency, which was more pronounced in animals exposed to the GA vapor. All animals recovered quickly and were less affected upon subsequent exposures. No clear dose-related differences were observed between groups which received the low versus the high GA concentration by inhalation. Glutaraldehyde was negative in the respiratory LLNA test. Exposure to GA by inhalation did not induce macroscopic enlargement of respiratory tract draining lymph nodes and inhalation exposure to GA did not induce significant increases in cytokine production in mandibular lymph nodes.

The Dow Chemical Company (1994) exposed groups of mice (six) dermally with 50 µL of glutaraldehyde. Seven days later, the animals were dosed again at half the first concentration applied to the dorsum of both ears. Fourteen days later, mice were sacrificed, and blood was collected and serum prepared. The concentration of serum IgE (measured using a sandwich ELISA) was compared to the responses of the positive control (trimellitic anhydride) and negative control (DNCB). Results were expressed as serum IgE concentration (µg/mL). Exposure of mice to the test material resulted in a dose-dependent increase in serum IgE relative to the control, elicitating mean values of 1.280 ± 0.193 µg/mL. A substantial increase in serum IgE levels relative to the historical control is considered indicative of the potential to cause sensitization of the respiratory tract. The data leads to the conclusion that glutaraldehyde exhibits some capacity for sensitization of the respiratory tract.

Human data

Several human population studies were identified regarding respiratory sensitization effects after exposure to the test substance.

Di Stefano et (1999) described a series of 24 health-care workers with respiratory symptoms suggestive of occupational asthma due to glutaraldehyde exposure. The history of asthmatic symptoms was investigated with peak expiratory flow rate (PEFR) monitoring, and in eight of the subjects, the specific bronchial provocation test (SBPT) was applied as reference standard for diagnosis of occupational asthma. Levels of glutaraldehyde were monitored in the challenge chamber during the SBPT. Specific IgE antibodies to glutaraldehyde were measured with a series of glutaraldehyde modified proteins. In the 8 workers who underwent SBPT, the diagnosis of occupational asthma was confirmed by a positive reaction. The mean level of glutaraldehyde observed during the challenge tests was 0.075 mg/m3. In 13 out of the 16 remaining workers, the serial PEFR monitoring showed a work-related effect. In 3 workers, there was no physiological confirmation of occupational asthma. Measurements of specific IgE antibodies to glutaraldehyde-modified proteins were positive in seven patients (29.1%).

After structured interviews Pechter et al (2005) found that the number and percentage of work-related asthma cases among health care workers for glutaraldehyde was 27 (N=305) cases, 9% respectively. In a study by McDonald et al (2000), chest and occupational physicians voluntarily reported 30, 128, and 133 cases of occupational asthma after exposure to glutaraldehyde for the periods 1989 -91, 1992 -94, and 1995 -97, respectively.

To gain insight in the mechanisms of glutaraldehyde activity concerning asthma, Palczynski et al (2001) performed a single-blind, placebo-controlled study with 11 health workers with occupational asthma and rhinitis due to glutaraldehyde. The control groups comprised 10 atopic subjects with perennial asthma and rhinitis and 10 healthy ones. There was a significant increase in eosinophile number and percentage, and albumin, eosinophile cationic protein, and tryptase concentrations in nasal lavage fluid from patients with occupational asthma and rhinitis due to glutaraldehyde when compared to controls. In a later study Palczynski et al (2005) evaluated bronchoalveolar lavage fluid (BALF) components and Clara cell protein (CC16) concentration in serum and BALF in patients with glutaraldehyde-induced asthma, before and after a specific inhalatory provocation test (SIPT) with glutaraldehyde, in comparison to atopic asthmatics and healthy individuals. In glutaraldehyde-sensitized asthmatics, the level of CC16 in BALF and serum was significantly lower at 24 h after SIPT in comparison with the values recorded prior to the experiment. In addition, there was a significant increase in the proportion of eosinophils, basophils und lymphocytes in BALF of glutaraldehyde-sensitized asthmatics obtained after SIPT.

In addition to the studies summarized above, Sutton et al (2007), a review by Takigawa et al (2006), a review by the Dutch Expert committee (2005), and Jachuk et al (1989) have published studies confirming the potential of occupational asthma or other respiratory sensitization effects as a result of glutaraldehyde exposures.

In contrast to the above, in an epidemiological study also from the Union Carbide Corporation (1993, later published by Teta et al 1995) the incidence of skin sensitization, respiratory sensitization and allergic blepharoconjunctivi was examined in 218 men assigned to glutaraldehyde production or drumming from its start-up in 1959 through 1992, at a chemical plant in West Virginia. Plant medical records, work restrictions, and clinic visits for accidental exposures were reviewed. There were no indications of glutaraldehyde-induced skin or respiratory sensitization, or allergic blepharoconjunctivitis in workers exposed at or below 0.2 ppm of glutaraldehyde. Also Pianiello et al (1997) found no evidence of increased incidence of respiratory effects, including asthmatic symptoms, in 135 nurses in 26 South Australian hospitals exposed to glutaraldehyde. Personal inhalational exposures were however generally low (overall geometric mean = 0.032 ppm).

Clinical cases:

Quirce et al (1999) described a case of a 61-year-old nurse working in a renal dialysis unit showing symptoms of glutaraldehyde induced asthma. The nurse developed symptoms of irritation of the eyes and upper respiratory tract, dyspnea on exertion, dry cough, and episodic attacks of wheezing. Skin prick test to 2 % glutaraldehyde was negative. Specific bronchial challenge test with activated 2 % glutaraldehyde showed no changes in FEV1 during a 24-h monitoring period. One week later the challenge test with 2 % glutaraldehyde was repeated, and it elicited an early asthmatic response. Although no late reaction was observed, a recurrent nocturnal asthmatic reaction occurred in the following days. The same test performed in two unexposed asthmatic patients elicited no response. Nicewicz et al (1986) described a case of a 25-year-old respiratory therapy technician. The therapy technician developed asthma following a period of relatively symptomless exposure to glutaraldehyde solution which was used in the cleaning of respiratory therapy equipment. On lung function testing, the woman demonstrated a delayed obstructive response after exposure to the chemical. Serum IgG and IgE levels as well as a scratch test performed with 2% glutaraldehyde solution were normal. The absolute eosinophile count was elevated. The woman changed positions within her department to reduce glutaraldehyde exposure, but upon re-exposure, she developed severe life-threatening status asthmaticus. At that time, the eosinophile count and immunoglobulins were normal. Ong et al (2004) described the case of a 32-year-old laboratory technician presented with adult-onset asthma 2 years after daily exposure to glutaraldehyde which was used to sterilise the mouthpieces used for lung function testing. A specific inhalational challenge test showed a 25% drop in forced expiratory volume in one second after exposure to glutaraldehyde but not after to a control substance. Corrado et al (1986) described the case of 4 nurses working in endoscopy units who complained of respiratory symptoms on exposure to alkaline glutaraldehyde used as a disinfectant for fibre-optic endoscopes. A provocation test was positive in two of these patients. Gannon et al (1995) described the cases of 8 workers who were referred for investigation of suspected occupational asthma following direct or indirect exposure to glutaraldehyde at work. The diagnosis of occupational asthma was confirmed in seven workers, all of whom had PEF records suggestive of occupational asthma and positive specific bronchial challenge tests to glutaraldehyde. Bronchial provocation testing was negative in one worker who was no longer exposed and who had a less clear-cut history of occupational asthma. Three workers also had a positive specific bronchial challenge to formaldehyde. Stenton et al (1994) described the case of a 46-year-old endoscopy nurse who developed symptoms suggestive of occupational asthma after 7 years of exposure to glutaraldehyde. An initial inhalation challenge test at the endoscopy suite caused a very dramatic immediate fall in FEV1 (1-s forced expiratory volume). A second series of double-blind challenges with controlled exposures to up to 0.32 ppm for 10 min) showed no obvious asthmatic reactions, in contrast to the results of the unblinded workplace challenge. Galdi et al (2005) described the case of a 47-year-old nurse who was exposed to glutaraldehyde while sterilizing devices for transesophageal echocardiography. Within a few minutes she suffered from sore throat, burning eyes, and dry cough. Later she also developed dyspnea and wheezing. In addition, she developed vocal cord dysfunction after exposure to glutaraldehyde. She had a history suggestive of reactive airway dysfunction syndrome (RADS).

Short description of key information:
In 2005, a review on glutaraldehyde data and health-based recommendations for occupational exposure limit was published by the Dutch Expert Committee on Occupational Standards of the Health Council of the Netherlands. In the review, case reports on occupational allergic contact dermatitis and positive patch test results from health-care workers were mentioned, which confirmed the skin sensitising potential of glutaraldehyde. Referring to the respiratory tract sensitizing potential of glutaraldehyde, it was reported that glutaraldehyde can cause asthmatic symptoms, thus, glutaraldehyde should be considered as a respiratory sensitizer.

Justification for classification or non-classification

According to Annex VI of EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008) and based on the available data, glutaraldehyde has to be classified as Skin Sens. 1A: H317 and Resp. Sens. 1: H334.