Lipase, triacylglycerol

Endpoint summary

Description of key information

Acute toxicity via the oral route has been tested. No signs of toxicity were observed in ten rats treated with a single oral dose of 2740 mg enzyme concentrate dry matter/kg bw (equivalent to 1940 mg active enzyme protein/kg bw). The test was conducted according to OECD guidelines and GLP standards.

Acute toxicity via the inhalation and dermal route is waived based on exposure considerations and the known properties of the substance.

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records

Reference

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From February 03, 1997 to August 08, 1997

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 420 (Acute Oral Toxicity - Fixed Dose Method)

Version / remarks:

and EEC directive 92/69/EEC.

Deviations:

no

GLP compliance:

yes

Test type:

fixed dose procedure

Limit test:

yes

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

- Source: Møllegaard Breeder, Ejby, Denmark
- Fasting period before dosing: overnight fasting prior to dosing
- Housing: Five of the same sex in transparent plastic boxes (Type IV 590 MAK overall dimensions 59 x 38 x 20 cm) with wire grid tops and aspen wood chips bedding. In a barrier maintained animal room with control of temperature and humidity
- Weight at the end of the akklimatization period: Males 154-156 g, females 124-131 g.
- Diet: Altromin rat/mouse Breeding 1320 diet pellets ad libitum
- Water: Water ad libitum
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22±3°C
- Humidity (%): 30-70%
- Air changes (per hr): 8-12
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 12 February 1997 to 26 Februay 1997

Route of administration:

oral: gavage

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

Undiluted test material

Doses:

Dose volume was 20 mL/kg bw, corresponding to a dose of 2740 mg enzyme concentrate dry matter/kg bw.

No. of animals per sex per dose:

5

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Observations for clinical signs of effect: before dosing, twice after dosing (day 1) and thereafter once per day for the following 14 days. Weighing immediately before dosing at day 1, day 8 and Day 15.
- Necropsy of survivors performed: yes

Statistics:

No

Sex:

male/female

Dose descriptor:

other: Fixed dose method - no effects were seen

Effect level:

20 mL/kg bw

Based on:

test mat.

Key result

Sex:

male/female

Dose descriptor:

other: Fixed dose method - no effects were seen

Effect level:

> 2 740 mg/kg bw

Based on:

other: Enzyme concentrate dry matter

Mortality:

No animals died during the study.

Clinical signs:

other: No clinical signs were observed in the dose group.

Gross pathology:

Effects on organs: No macroscopic alterations were observed at the terminal post mortem examination.

Interpretation of results:

GHS criteria not met

Conclusions:

No signs of toxicity were observed among the rats treated with a single oral dose of 2740 mg enzyme concentrate dry matter/kg bw (equivalent to 1940 mg active enzyme protein/kg bw), which was the highest possible dose at dose volume 20 mL/kg, using the undiluted test item.

Executive summary:

The study was conducted in accordance with the OECD Guideline No 420, "Acute Oral Toxicity - Fixed dose Method" and EEC directive 92/69/EEC.

The test item was supplied as a brown liquid ready to use. The dose volume administered was 20 mL of the undiluted test material per kg body weight, corresponding to a dosage of

2.74 g enzyme concentrate dry matter/kg bw. A group of five male and five female rats received the test material by a single oral administration (gavage). The animals were subjected to clinical observations daily for a fourteen day observation period and at termination of the study, gross necropsy of all animals was carried out.

No clinical effects were observed and the overall body weight gain during the study was considered to be normal. The post-mortem inspection revealed no abnormalities.

In conclusion, no signs of toxicity were observed among the rats treated with a single oral dose of 2740 mg enzyme concentrate dry matter/kg bw (equivalent to 1940 mg active enzyme protein/kg bw).

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Quality of whole database:

Toxicological data has been generated within the enzyme producing industry during the last 40 years. Substantial documentation on the safety of the production strains have been generated, and the enzyme test materials are thoroughly characterized. High quality studies for all relevant endpoints, in vivo studies as well as in vitro studies, show that industrial enzymes from well-known and well-characterized production strains have very similar safety profiles across the catalytic activities. Read-across can therefore be applied for the majority of toxicological endpoints. The database can thus be considered of high quality.

Acute toxicity: via inhalation route

Link to relevant study records

Reference

Endpoint:

acute toxicity: inhalation

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

other:

Endpoint conclusion

Endpoint conclusion:

no study available

Quality of whole database:

Toxicological data has been generated within the enzyme producing industry during the last 40 years. Substantial documentation on the safety of the production strains have been generated, and the enzyme test materials are thoroughly characterized. High quality studies for all relevant endpoints, in vivo studies as well as in vitro studies, show that industrial enzymes from well-known and well-characterized production strains have very similar safety profiles across the catalytic activities. Read-across can therefore be applied for the majority of toxicological endpoints. The database can thus be considered of high quality.

Acute toxicity: via dermal route

Link to relevant study records

Reference

Endpoint:

acute toxicity: dermal

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

the study does not need to be conducted because the physicochemical and toxicological properties suggest no potential for a significant rate of absorption through the skin

Endpoint conclusion

Endpoint conclusion:

no study available

Quality of whole database:

Toxicological data has been generated within the enzyme producing industry during the last 40 years. Substantial documentation on the safety of the production strains have been generated, and the enzyme test materials are thoroughly characterized. High quality studies for all relevant endpoints, in vivo studies as well as in vitro studies, show that industrial enzymes from well-known and well-characterized production strains have very similar safety profiles across the catalytic activities. Read-across can therefore be applied for the majority of toxicological endpoints. The database can thus be considered of high quality.

Additional information

In general, enzymes are of very low toxicity due to ready biodegradability and very low bioavailability. In traditional acute toxicity testing, mortality has been the endpoint. However, because enzymes show very low toxicity, extremely high doses that are far above human exposure levels typically have been applied. Therefore, acute toxicity studies are not considered to provide appropriate knowledge and are as such not a relevant test system for enzymes. Systemic exposure by the dermal route is unlikely based on the existing toxicokinetic knowledge of enzymes, which due to their relatively large molecular weight, are not expected to be absorbed through the skin (Basketter et al. 2008, Smith Pease et al. 2002). Therefore, it can be safely assumed that technical enzymes do not exert any acute dermal toxicity (Basketter et al 2012). This conclusion is confirmed by the toxicological data available. Sub-acute dermal toxicity studies with protease in rabbits (Novozymes, unpublished data) did not provide evidence for systemic effect to enzymes. This finding is confirmed by data from acute dermal toxicity studies (Novozymes, unpublished data) of other enzyme products in both rats and rabbits. None of these studies revealed any acute toxic effect through the dermal administration route. No clinical signs or adverse effects due to systemic exposure could be observed. Data waivers will further be established through exposure scenarios, i.e. no significant dermal exposure to consumers and professionals due to the toxicologically insignificant enzyme concentrations in end products and in the case of workers due to occupational hygiene measures associated with the prevention of respiratory allergy which includes protective clothing. In conclusion, toxicokinetic data together with evidence from animal studies and historical human experience derived from the use of detergent enzymes for decades confirm that exposure to technical enzymes will not result in any toxicologically relevant uptake by dermal route. Acute systemic exposure to a toxicologically significant amount of enzymes by this route can, therefore, be excluded and will further be prohibited by the obligatory setting of a DMEL value for enzymes, resulting in negligible exposure to enzymes (Basketter et al 2010). In vivo acute dermal toxicity studies will not add any value and cannot be expected to provide valuable knowledge and are considered scientifically and ethically unjustified. Therefore, in accordance with column 2 of REACH Annex VIII acute toxicity testing by the dermal route is inappropriate.  

References:

- Basketter DA, English JS, Wakelin SH, White IR (2008). Enzymes, detergents and skin: facts and fantasies.Br. J. Dermatol., 158 (6):1177-1181.

- Smith Pease CK, White IR, Basketter DA (2002). Skin as a route of exposure to protein allergens.Clin. Exp. Dermatol., 27(4):296-300.  

- Basketter D, Berg N, Broekhuizen C, Fieldsend M, Kirkwood S, Kluin C, Mathieu S, Rodriguez C (2012a). Enzymes in Cleaning Products: An Overview of Toxicological Properties and Risk Assessment/Management.Regul. Toxicol. Pharmacol., 64(1):117-123.

- Basketter DA, Broekhuizen C, Fieldsend M, Kirkwood S, Mascarenhas R, Maurer K, Pedersen C, Rodriguez C, Schiff HE (2010). Defining occupational and consumer exposure limits for enzyme protein respiratory allergens under REACH.Toxicology, 268(3):165-170.

Justification for classification or non-classification

Based on the low acute oral toxicity of lipase, the low likelihood of absorption of enzymes through the skin due to the physico-chemical properties of the enzyme and the low exposure to enzymes by inhalation enforced by the respiratory allergy exposure limits, lipase should not be classified.