4-methylpentan-2-one oxime

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

14 November 1989 - 06 December 1989

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1990

Report date:

1990

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Deviations:

no

GLP compliance:

yes

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

The objective of the study was to assess the potential of MIBKO to elicit an allergic contact sensitization response in guinea pigs. The Local Lymph node Assay is recognized as the preferred test method. However, this in vivo guinea pig maximization test was performed prior to enforcement of the OECD guideline 429 ; and was deemed suited for skin sensitisation potential assessment of the test substance.

Test material

Test material information

Constituent 1

Reference substance name:

4-methylpentan-2-one oxime

EC Number:

203-298-2

EC Name:

4-methylpentan-2-one oxime

Cas Number:

105-44-2

Molecular formula:

C6H13NO

IUPAC Name:

4-methylpentan-2-one oxime

Test material form:

liquid

Specific details on test material used for the study:

- Test article code Number: MIBKO (199-89B)
- Physical description: clear liquid
- Lot number: 37905-27-4
- Purity: 100%
- Physicochemical parameters: density 0.88 g/ml; solubility 0.9% in water
- Expiration date: No data
- Storage conditions: 65-85°F

In vivo test system

Test animals

Species:

guinea pig

Strain:

Hartley

Remarks:

Albino

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: CAMM Research Laboratory Animals Wayne, New Jersey 07470 , USA
- Age at study initiation: Young adult
- Weight at study initiation: 275 - 425 g
- Housing: Individually housed in stainless steel, wire mesh bottom cages.
- Diet: ad libitum , Fresh Aqway Certified Guinea Pig - Feed was provided throuqhout both the acclimation and scudy periods.
- Water: ad libitum, via an automatic watering system
- Acclimatization period: 8 days

ENVIRONMENTAL CONDITIONS
- Temperature: 64 - 79 °C
- Humidity: 40 - 70 %
- Photoperiod: 12 hrs dark / 12 hrs light

IN-LIFE DATES: From 07 Nov 1989 To: 15 Dec 1989

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal

Vehicle:

propylene glycol

Concentration / amount:

5.0% w/v

Day(s)/duration:

Day 1 of treatment

Induction 2

Route:

intradermal

Vehicle:

other: Freunds Complete Adjuvant (FCA) emulsion

Concentration / amount:

5.0% w/v

Day(s)/duration:

Day 1 of treatment

Induction 3

Route:

epicutaneous, occlusive

Vehicle:

propylene glycol

Concentration / amount:

100% w/v

Day(s)/duration:

Day 7 of treatment

Challenge

Challenge 1

No.:

#1

Route:

epicutaneous, occlusive

Vehicle:

propylene glycol

Concentration / amount:

5% w/v of test article for negative control and test animals
0.1% w/v of Dinitrochlorobenzene (DNCB) to positive control animals

Day(s)/duration:

Day 21 of the study

Challenge 2

No.:

#2

Route:

epicutaneous, occlusive

Vehicle:

propylene glycol

Concentration / amount:

5% w/v of the test item to the negative control and test animals

Day(s)/duration:

Day 28 of the study

No. of animals per dose:

MIBKO
Intradermal Range Finding : 4 animals
Topical Range Finding : 8 animals
Induction/Challenge : 15 animals
Challenge control : 5 animals
Rechallenge control : 5 animals

DNCB
Induction/Challenge : 10 animals
Challenge Control : 5 animals

TOTAL: 52 animals

Details on study design:

BODY WEIGHTS
Body weights were measured on Days -7, 0, 7, 14, 21 and 28 of the study for all animals used on the main study. Body weights for the range finding animals were measured on the day of dosing only.

INTRADERMAL RANGE FINDING TEST
Four w/v concentrations of the test article were prepared in Propylene Glycol and also in Freund's Complete Adjuvant (FCA) Emulsion (1:1 FCA in sterile water for injection, USP). The following concentrations were used: 0.1%, 1.0%, 3.0% and 5.0%. On the day of dosing, the hair was removed from the backs of four guinea pigs using a small animal clipper and a size No. 40 blade.
Two pair of intradermal injections at a volume of 0.1 ml each were made along the spine on each of the four guinea pigs (two injections per side)
Approximately 24 and 48 hours following injection, all sites were scored for erythema.

TOPICAL RANGE FINDING STUDIES
Four w/v concentrations of the test article were prepared in propylene glycol. A second topical range finding study was performed due to the irritating results noted following the first range finding study.
- No. of exposures: 1 (topical application)
- Type of epicutaneous induction: occlusive
- Exposure period: on D7 (epidermal, 48-hr exposure); approximately 24 hours following patch application, all wrappings were removed and any residual test article was wiped and washed with a gauze pad moistened with propylene glycol. Approximately 21 hours following patch removal, the test sites were reclipped as previously described and three hours thereafter the test sites were scored for irritation. The scoring was repeated 24 hours later.
- Test groups:
Study 1: The test item at 100%, 50%, 25%, and 10%, in propylene Glycol
Study 2: The test item at 10%, 5%, and 1.0%, in propylene Glycol
- Site: on the backs of the animals, two pads on each side of the spine with one near the shoulder and one near the hip. The animals were then occlusively wrapped with a sheet of nonreactive, rubberized material which was securely taped in place with nonirritating tape. This covering was secured with adhesive Blenderm tape.
- Frequency of applications: once
- Duration: 48 hours
- a volume of 0.4 mL of each concentration of the test article was applied individually to 2x2 cm Webril pads.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 2 (intradermal injections + topical application)
- Type of epicutaneous induction: occlusive
- Exposure period: on D0 (intradermal) and D7 (epidermal, 48-hr exposure)
- Test groups:
* INTRADERMAL INDUCTION
- Site: on the day of dosing , the hair was clipped from the scapular region of the test, positive and negative control animals. Three pairs of intradermal injections were made in the clipped area of each animal. The injections were kept within a boundary of a 2x4cm area with one row of 3 injections on each side of the spine:
1) 0.1mL of FCA emulsion
2) 0.1 mL of 5% (w/w) test article in propylene glycol
3) 0.1 mL of 5% (w/w) test article
- positive control animals:
1) 0.1mL of FCA emulsion
2) 0.1mL of 0.1% DNCB in PG
3) 0.1mL of 0.1% DNCB in FCA emulsion
- negative control animals
1) 0.1mL of FCA emulsion
2) 0.1mL of PG
3) 0.1mL of 5% PG in FCA emulsion
Injections 1 and 2 were administered close to one another and nearest to the head, injection 3 was administered most caudally.

*TOPICAL ADMINISTRATION (day 7)
- on the day of dosing, the hair was reclipped from the scapular region of the test, positive and negative control animals.
- Control group: positive control 0.8mL 0.1% DNCB and negative control 0.8mL propylene glycol
- Test group: 0.8mL test item (neat) at 100% (w/v)
- Site: Each dose was first applied to a 2 x 4 cm Webril pad which was immediately applied over the intradermal injection sites. Immediately following patch application, the sites were wrapped.
- Frequency of applications: once
- Duration: 48 hours, approx 48hrs after application, all wrappings were removed

B. CHALLENGE EXPOSURE
- on day 28, the hair was clipped from the left and right flanks of the test, positive and negative control animals
- No. of exposures: 1 (topical application)
- Day of challenge: day 28
- Exposure period: 24 hours
- Test groups and negative challenge control: 0.4mL of 5% w/v test article in Propelyne Glycol on the posterior left flank and 0.4 mL Propylene Glycol only on the posterior right flank.
-Positive control group (incl. positive challenge group): 0.4 mL of 0.1% DNCB in Propylene Glycol on the posterior left flank and0.4mL of propylene Glycol on the posterior right flank
- each dose was first applied to a 2x2 cm webril pad which was immediately applied to the previously designated sites. Immediately following patch application, the animals were wrapped.
- Evaluation: approx 24 hours following patch application, all wrappings were removed and, if necessary, any residual material was wiped and washed from the site using a gauze pad moistened with PG. Approx. 21 hours following patch removal, all test sites were reclipped and three hours thereafter, the sites were scored for irritation. The scoring was repeated 24 hours later (approx 48 hours post patch removal).

C. RECHALLENGE PHASE
- No. of exposures: 1 (topical application)
- Day of challenge: day 28, the hair was clipped from the left and the right flanks of the test and negative rechallenge control group animals.
-Test groups and negative rechallenge control: 0.4 mL of 5% w/v test article in Propelyne Glycol on the anterior left flank and 0.4mL of Propylene Glycol on the anterior right flank.
- each dose was first applied to a 2x2cm Webril pad which was immediately applied to the designated sites.
- Evaluation: 48 hours post patch removal

Challenge controls:

- Negative challenge control: 5% w/v test article in Propylene Glycol on the posterior left flank and Propylene Glycol only on the posterior right flank.
- Positive challenge group: 0.1% DNCB in Propylene Glycol on the posterior left flank and propylene Glycol on the posterior right flank

Positive control substance(s):

yes

Remarks:

Dinitrochlorobenzene (DNCB) (CAS No: ‎97-00-7)

Results and discussion

Positive control results:

At challenge, all of the DNCB positive control animals had positive scores of one or greater at both 24 and 48 hour scoring periods. None of the DNCB challenge control animals had positive responses.
Under the conditions of this test, the positive control DNCB was considered to be an Extreme Sensitizer (100% sensitized) and test article MIBKO (199-898) was considered to be a Nonsensitizer (0% sensitized).

In vivo (non-LLNA)

Results

Results 1

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

5%

No. with + reactions:

1

Total no. in group:

15

Remarks on result:

other: score was +/-

Results 2

Reading:

1st reading

Hours after challenge:

48

Group:

test chemical

Dose level:

5%

No. with + reactions:

0

Total no. in group:

15

Remarks on result:

no indication of skin sensitisation

Results 3

Reading:

rechallenge

Hours after challenge:

24

Group:

test chemical

Dose level:

5%

No. with + reactions:

2

Total no. in group:

15

Remarks on result:

other: score was +/-

Results 4

Reading:

rechallenge

Hours after challenge:

48

Group:

test chemical

Dose level:

5%

No. with + reactions:

2

Total no. in group:

15

Remarks on result:

other: score was +/-

Results 5

Reading:

1st reading

Group:

negative control

Dose level:

0%; propylene glycol

No. with + reactions:

0

Total no. in group:

15

Remarks on result:

no indication of skin sensitisation

Results 6

Reading:

1st reading

Hours after challenge:

24

Group:

positive control

Dose level:

0.1% DNCB in PG

No. with + reactions:

15

Total no. in group:

15

Clinical observations:

eschar at test site of 1 animal

Remarks on result:

positive indication of skin sensitisation

Remarks:

confirmed as extreme sensitiser

Results 7

Reading:

1st reading

Hours after challenge:

48

Group:

positive control

Dose level:

0.1% DNCB in PG

No. with + reactions:

15

Total no. in group:

15

Clinical observations:

eschar at test site in 4/15 animals

Remarks on result:

positive indication of skin sensitisation

Remarks:

confirmed as extreme sensitiser

Any other information on results incl. tables

Body weight:

All animals gained weight by the end of the study period.

Mortality:

- One animal in the test article group died during the study period.

- A gross necropsy was performed on this animal : heart was discolored yellow and the stomach and the gastrointestinal tract were distended with gas. All other organs and tissues appeared normal.

Detailed results on animals are shown in the following tables:

CHALLENGE SCORES

Table 1: MIBKO challenge control

Animal ID #	24 H Erythema test	24 H Erythema vehicle	48H Erythema test	48H Erythema vehicle
Q821	0	0	0	0
Q824	0	0	0	0
Q858	0	0	0	0
Q862	0	0	0	0
Q892	0	0	0	0

None of the challenge control animals had positive responses.

Table 2: MIBKO test animals

Animal ID #	24 H Erythema test	24 H Erythema vehicle	48H Erythema test	48H Erythema vehicle
Q823	0	0	0	0
Q826	0	0	0	0
Q829	*	*	*	*
Q831	0	0	0	0
Q835	0	0	0	0
Q837	0	0	0	0
Q839	0	0	0	0
Q842	0	0	0	0
Q865	0	0	0	0
Q871	0	0	0	0
Q872	0	0	0	0
Q885	0	0	0	0
Q893	0	0	0	0
Q897	0	0	0	0
Q904	+/-	0	0	0

*Not Applicable, animal died on test.

One animal had a score of +/- at the 24 hour scoring period. this score had reduced to 0 at the 48 hour scoring period.

Table 3: DNCB Positive Control

Animal ID #	24 H Erythema test	24 H Erythema vehicle	48H Erythema test	48H Erythema vehicle
Q832	2	0	1	0
Q843	2	0	2	0
Q854	3	0	3	0
Q855	3	0	3**	0
Q860	3	0	2	0
Q864	2	0	3**	0
Q877	2	0	1	0
Q880	3**	0	3**	0
Q895	3	0	3	0
Q899	2	0	3**	0

** Eschar at test site

Table 4: DNCB Challenge Control

Animal ID #	24 H Erythema test	24 H Erythema vehicle	48H Erythema test	48H Erythema vehicle
Q850	0	0	0	0
Q853	0	0	0	0
Q857	0	0	0	0
Q869	0	0	0	0
Q902	0	0	0	0

RECHALLENGE SCORES

Table 5: MIBKO Rechallenge control

Animal ID #	24 H Erythema	48 H Erythema
Q833	0	0
Q838	0	0
Q881	0	0
Q886	0	0
Q900	0	0

 Animal ID #  24 H Erythema   48 H Erythema

Table 6 : MIBKO Test Animals

Animal ID #	24 H Erythema	48 H Erythema
Q823	0	0
Q826	0	0
Q829	*	*
Q831	0	0
Q835	0	0
Q837	0	0
Q839	0	0
Q842	0	0
Q865	0	0
Q871	+/-	+/-
Q872	0	0
Q885	0	0
Q893	0	0
Q897	0	0
Q904	+/-	+/-

At rechallenge, two of the test animals had scores of +/- at both the 24 and 48 hour scoring periods. These responses of +/- are considered to be equivocal and therefore not considered to be positive sensitising responses.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Under the conditions of this test, the positive control DNCB was considered to be an Extreme Sensitizer (100% sensitized) and test article MIBKO (199-89B) was considered to be a Nonsensitizer (0% sensitized).

Executive summary:

MIBKO (Sample No 199 -89B) was evaluated for allergic contact sensitization using the maximization test on female guinea pigs. This involved two induction exposures. MIBKO was first administered by intradermal injection and second by topical exposure, followed by a topical challenge exposure 21 days after the first induction. At challenge, none of the animals developed any skin reactions indicating sensitisation. A topical rechallenge phase was performed on day 28 to confirm the negative results of the challenge phase. MIBKO produced no signs of contact sensitization and was classified as a nonsensitizer (0% sensitized).