1,2-dihydro-3H-1,2,4-triazol-3-one

Dermal absorption

Administrative data

Endpoint:

dermal absorption in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

22nd December 2011to 17th July 2012

Reliability:

1 (reliable without restriction)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 428 (Skin Absorption: In Vitro Method)

Test material

Test material information

Constituent 1

Reference substance name:

NTO

IUPAC Name:

NTO

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Details on test material:

- Name of test material (as cited in study report): 3-nitro-1,2,4-triazole-5-one (NTO)
- Physical state: a white powder

Radiolabelling:

no

Test animals

Species:

other: Full thickness abdominal human skin from autopsy or surgery

Administration / exposure

Type of coverage:

open

Vehicle:

physiological saline

Duration of exposure:

24 hours

Doses:

Infinite dose of 100 mg

No. of animals per group:

5 cells

Control animals:

no

Details on in vitro test system (if applicable):

Full thickness abdominal human skin (from autopsy or surgery) was obtained from the National Disease Research Interchange (NDRI), Philadelphia, Pennsylvania. The whole skin was stored frozen at -85 degrees Centigrade (°C). Before starting the experiments, the skin was placed in a Petri dish containing a buffer solution at room temperature. The subcutaneous fat was removed with blunt forceps, and the skin was cleaned with a buffer solution and placed on a dissecting board. Circular pieces of skin sections (0.64 cm2) were prepared from the skin with the help of a metallic Arch Punch (18 mm). The epidermis was teased from dermis as described previously (Frasch et al. 2011). The skin discs were wrapped in Saran® Wrap and submerged in a 60°C buffer for approximately 70 seconds. The skin was unwrapped and placed in a Petri dish containing a buffered solution. The dermis of the skin discs was teased from the epidermis carefully using blunt forceps or cotton swabs under a binocular microscope. Care was taken not to damage the skin. During epidermal membrane separation, a thin layer of stratum corneum was easily separated from the skin. The epidermal membranes remaining were used for testing. The epidermal membranes were evaluated under a dissection microscope to ensure that they were free of any damage. The epidermal membranes (without stratum corneum) were placed on wax paper moistened with a buffered solution and frozen (-30°C) prior to experimental use. Before use, the stored epidermal membrane discs were thawed at room temperature and checked again for any damage under the dissection microscope before testing.
The in vitro static diffusion cell system was composed of water-jacketed glass cells with a receptor chamber volume of 5 milliliters (mL). The individual cells were coupled to a water distribution system, which was connected to a recirculating water bath adjusted to allow the skin, mounted on the in vitro cells, to be maintained at 32°C. Single Franz diffusion cells were mounted in a rack which can hold six cells. The exposed skin surface was 0.64 cm2. The receptor chamber was filled with 0.9% phosphate buffered saline (PBS) and the skin disk was placed above. The donor chamber was clamped over the skin disk on the receptor chamber. The donor chamber is equipped with a side arm which allows for the collection of samples from the receptor fluid for the measurement of penetrated chemicals. The system was allowed to equilibrate before measuring skin integrity.
Individual chemical samples of powdered material were applied to the moistened human epidermal membranes in the donor chamber at an infinite dose (100 mg).At different times, 0.1 mL of receptor fluid was collected with a syringe and was placed in a vial containing acetonitrile (0.9 mL) for further dilution and analysis by HPLC. The volume of the receptor fluid was maintained at 5 mL by replenishing it with 0.1 mL of buffer after each sample collection. Samples were collected at 1, 2, 4, 6, 8, and 24 hours for NTO. The steady state flux of NTO was between 6 to 8 hours.

Results and discussion

Signs and symptoms of toxicity:

not examined

Dermal irritation:

not examined

Absorption in different matrices:

The steady state of absorption was determined to be from 6-8 hours and did not change thereafter so the penetration studies were studied at 8 hours for all chemicals. The dermal absorption rate for NTO at 8 hours was 338.2 μg/ cm2 /hr. The cumulative dermal absorption of NTO was found to be 5843 μg/ cm2, while at 24 hours the cumulative absorption was 4250 μg/ cm2

Total recovery:

N/a

Percutaneous absorption

Time point:

8 h

Parameter:

percentage

Absorption:

0.528 %

Applicant's summary and conclusion

Conclusions:

The analysis of absorbed chemical in the receptor fluid data showed that the steady state flux to an infinite dose (100 mg) of neat 3-Nitro-1,2,4-triazol-5-one (NTO) was 338.2 μg/ cm2 /hr
In an in vitro dermal absorption study ( U.S. Army Public Health Command, 2012) the penetration rate of 3-Nitro-1,2,4-triazol-5-one (NTO) was found to be 338.2 µg/cm2/hr for an administered dose of 100 mg. Considering penetration of skin area of 0.64 cm2, the penetration rate is 1.72 µg/hr (338.2 µg/cm2/hr ÷ 0.64 cm2 = 528.4375 µg/hr) .

From this a dermal absorption rate for NTO (% total absorbed) is 0.528% (=528.4375 µg /100 mg).

Executive summary:

The in vitro dermal penetration of 3-nitro-1,2,4-triazol-5-one (NTO) in static Franz diffusion cells was studied. In this system, the human epidermal membranes were prepared from frozen cadaver skin and were mounted on static diffusion receptor cells so that the visceral side was intact with the receptor fluid. The test chemical at infinite dose (100 milligrams (mg)) powder was carefully placed on the mounted skin in the donor chamber, and at different times (1, 2, 4, 6 and 8 hours) 0.1 milliliter (mL) of receptor fluid (buffer) was collected and quantified by High Performance Liquid Chromatography. The penetration rate was calculated in micrograms per square meter per hour (μg/cm2/hr). The analysis of absorbed chemical in the receptor fluid data showed that steady state of flux of infinite dose of neat NTO was 338.2 μg/cm2/hr.

Using the measured penetration rate of NTO of 338.2µg/cm2/hr for an administered dose of 100 mg and considering penetration of skin area of 0.64 cm2, the penetration rate is 528.4375 µg/hr (338.2 µg/cm2/hr ÷ 0.64 cm2 = 528.4375 µg/hr) .From this a dermal absorption rate (% total absorbed) is 0.528% (=528.4375 µg /100 mg) for NTO can be calculated.