Reaction mass of ethanol and propan-2-ol

Additional toxicological data

Administrative data

Endpoint:

additional toxicological information

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Not a guideline study and report does not state whether performed according to GLP. However, data appears well documented and scientifically acceptable

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Title:

Model studies for evaluating the acute neurobehavioural effects of complex hydrocarbon solvents I. Validation of methods with ethanol

Author:

McKee RH, Lammers JHCM, Hoogendijk EMG, Emmen HH, Muijser H, Barsotti DA, Owen DE & Kulig BM

Year:

2006

Bibliographic source:

Neurotoxicology 27, 1064-1079

Materials and methods

Type of study / information:

Behavioural tests following acute ethanol exposures were undertaken in an attempt to evaluate various measures of cognitive and psychomotor functioning in rats compared with human volunteers, and to assess relationships between internal levels of exposure and nervous system function.

Principles of method if other than guideline:

A range of tests including standardised functional observational measures, spontaneous motor activity assessments, and learned visual discrimination performance were undertaken in rats to evaluate acute central nervous system effects. In humans, several behavioural assessments (including tests of mood and affect, psychomotor skills, attention, perceptual coding, learning and memory) were performed. Blood ethanol levels of both rats and humans were taken to verify internal dose.

GLP compliance:

not specified

Test material

Test material information

Constituent 1

Reference substance name:

Ethanol

EC Number:

200-578-6

EC Name:

Ethanol

Cas Number:

64-17-5

Molecular formula:

C2H6O

IUPAC Name:

ethanol

Details on test material:

Described as potable ethanol of agricultural origin with an ethanol content of 96.1%. Because of manufacturing methods commonly used, most of the remainder is likely to be water.

Results and discussion

Any other information on results incl. tables

Animal tests.

No significant clinical signs or body weight changes were apparently observed.

Regarding the measures in the functional observational battery, statistically significant changes were seen in gait score, mean-foot splay (both measuring neuromuscular function), and tail-pinch response (sensori-motor function) at doses of 1 and 2 g ethanol/kg bw, and in mean fore-limb grip strength (neuromuscular function) and click response (sensori-motor function) at 2 g ethanol/kg bw only, compared with controls (see Table 2 for details).

Table 2. Results of functional observations in rats following ethanol administration

FOB measure	Dose (g/kg)	Pre-treatment test result (mean ± S.E.)	Test result (mean ± S.E.)
Gait score^a	0   0.5  1.0   2.0	1.00 ± 0.00  1.00 ± 0.00  1.00 ± 0.00  1.00 ± 0.00	1.00 ± 0.00 1.00 ± 0.00  2.00 ± 0.19*   3.63 ± 0.18**
Mean foot-splay^b(mm)	0   0.5  1.0   2.0	62.69 ± 3.40 65.94 ± 1.46 65.25 ± 3.52 57.81 ± 2.72	61.75 ± 1.43 68.88 ± 1.45  71.06 ± 4.13* 79.63 ± 3.89**
Mean fore-limb grip-strength^c(g)	0   0.5  1.0   2.0	1403 ± 62.97  1264 ± 96.28    1303 ± 130.81  1236 ± 82.30	856 ± 61.25    860 ± 71.22    971 ± 72.12    575 ± 69.47*
Click response^d	0   0.5  1.0   2.0	3.50 ± 0.19  3.50 ± 0.19  3.38 ± 0.26  3.63 ± 0.26	3.75 ± 0.15  3.50 ± 0.27  2.88 ± 0.35  2.50 ± 0.38*
Tail pinch response^e	0   0.5   1.0   2.0	3.75 ± 0.25  3.25 ± 0.37  3.50 ± 0.33  3.50 ± 0.33	4.00 ± 0.00  4.00 ± 0.00  3.00 ± 0.38*  2.00 ± 0.00**

*p<0.05; **p<0.01.

^aGait scored indicates the severity of gait changes and is scored as 1 (normal) to 4 (severely abnormal).

^bFoot-splay is the distance between the fourth digit footpads of a rat which is dropped from approximately 40 cm.

^cGrip-strength is measured with a strain gauge.

^dClick response was scored as 0 (no reaction) to 5 (exaggerated reaction).

^eTail pinch response was scored as 0 (no reaction) to 5 (exaggerated reaction).

No statistically significant changes between baseline and following ethanol exposure were seen in rats measured for arousal (excitation), mean hind-limb grip strength (neuromuscular function), and approach response or touch response (both measures of sensori-motor function).

With regards motor activity parameters, a statistically significant reduction in total distance transversed and number of movements initiated was seen at 2 g ethanol/kg bw compared to controls (see Table 3 for further details).

Table 3. Results of motor activity measurements in rats following ethanol administration

Parameter	Dose (g/kg)	Pre-treatment test result (mean ± S.E.)	Test result (mean ± S.E.)
Total distance traversed (cm)	0 0.5 1.0 2.0	3624 ± 200 3767 ± 328 3180 ± 223 3863 ± 261	3225 ± 144 2933 ± 294 2950 ± 270 1770 ± 236***
Number of movements initiated	0   0.5   1.0   2.0	54.2 ± 3.6 55.9 ± 5.1 57.0 ± 4.1 59.2 ± 3.2	52.4 ± 3.8 46.5 ± 5.6  36.8 ± 3.1*  19.9 ± 2.2***

*p<0.05; ***p<0.001.

No statistically significant differences were seen at lower doses for these parameters, or between ethanol exposed groups and controls at any dose regarding the mean velocity of locomotion.

Several measures of visual discrimination were assessed. No statistically significant differences were observed between ethanol exposed rats and controls in tests including percentage of reinforcements, discrimination ratio, total errors, repetitive errors, the percentage of inter-trial interval periods, the number of total and repetitive inter-trial interval periods, and number of lever response latencies greater than 6 seconds. A statistically significant reduction in the number of trials completed and lever response latencies (of a maximum of one or two seconds) and drink response latency was seen in the rats exposed to 2 g ethanol/kg bw, compared to controls (see Table 4 for further details). No statistically significant differences were seen for these measures of visual discrimination performance at doses of 1 g ethanol/kg bw and below, compared to controls.

Table 4. Mean group data for visual discrimination tests in rats

Parameter	Dose^a(g/kg)	Pre-treatment test results (mean ± S.E.)	Test results (mean ± S.E.)
Trials completed	0   0.5   1.0   2.0	100.00 ± 0.00 100.00 ± 0.00 100.00 ± 0.00 100.00 ± 0.00	100.00 ± 0.00 100.00 ± 0.00 98.38 ± 1.63 27.88 ±6.29**^b
Lever response latency (s)	0   0.5   1.0   2.0	2.01 ± 0.29 1.88 ± 0.20 2.23 ± 0.35 2.61 ± 0.72	2.46 ± 0.44 2.29 ± 0.35 4.40 ± 1.46     51.04 ± 23.38**
Number of lever response latencies <1 s	0   0.5   1.0   2.0	40.00 ± 6.28 44.13 ± 5.02 37.00 ± 4.06 43.13 ± 4.57	31.25 ± 5.14 35.63 ± 7.42 28.63 ± 3.69    2.29 ± 1.04**
Number of lever response latencies <2 s	0   0.5   1.0   2.0	72.38 ± 3.83 76.13 ± 2.41 71.75 ± 3.67 72.63 ± 2.08	72.25 ± 2.97 67.88 ± 4.75 66.38 ± 4.78   11.00 ± 2.79**
Drink response latency (s)	0  0.5  1.0   2.0	0.34 ± 0.02 0.32 ± 0.02 0.37 ± 0.02 0.37 ± 0.02	0.42 ± 0.03 0.37 ± 0.02 0.44 ± 0.02   0.79 ± 0.05**

^aThere were eight rats in each group.

^bThe statistical comparisons are to the corresponding control groups.

**p<0.01.

Mean blood ethanol levels in the 0.5, 1, and 2 g/kg bw groups were 0.458, 0.996 and 1.974 g/L, demonstrating a linear relationship between the amount of ethanol administered to the rats and the blood levels 20 minutes later.

Human volunteer tests.

For measures of mood and affect (including scales for depression, anger, fatigue, vigour, or tension) and for perceptual coding (involving a symbol-digit substitution test and a pattern comparison test), there were no statistically significant differences between volunteers exposed to ethanol or the placebo, assessed at 1 or 3 hr after consuming the beverages. For psychomotor skills, there were no differences in measures of hand-eye co-ordination (sinus and sawtooth condition) or in the (dominant and non-dominant hand) finger tapping tests, between ethanol exposed and placebo treated volunteers, tested at 1 and 3 hr post dosing. For the finger tapping test with alternating hands, performance was significantly lowered in volunteers exposed to ethanol when tested at 1 hr after dosing (but not at 3 hr post dosing), compared to placebo (see Table 5 for details). In the domain of attention, no statistically significant effects of treatment compared with placebo were detected in latency of responding in the simple reaction time test (assessed at 30 minutes, 1, 2 or 3 hr post dosing), or (as assessed at 1 and 3 hr post dosing) the colour word vigilance test and the switching attention tests (except for the "switching test" at 3 hr - see Table 5 for details).

Table 5. Summarized results of neurobehavioral tests in volunteer studies

Test	Test time point	Placebo		Ethanol
		Mean	S.E.M.	Mean	S.E.M.
Finger tapping test (# in 30 s)
Alternating hands	Pre-dosing 1 h 3 h	266 272 266	17 14 12	268  251* 276	13 14 16
Switching attention test (ms)
Switching	Pre-dosing 1 h 3 h	374 357 347	19 14 16	361 355  351*	15 17 17

*p<0.05.

There appeared to be a relationship between blood alcohol levels and performance in both the simple reaction time test and the colour word vigilance test, suggesting a relationship with treatment. For measures of learning and memory including the spatial memory span, digit memory span backwards and the pattern memory tests, no statistically significant treatment related effects were seen. In contrast, performance in the verbal memory tests (free and cued recall) were significantly affected when assessed at 1 hr compared to 3 hr-post ethanol treatment.

Mean blood ethanol levels peaked at about 0.53 g ethanol/L one hour after exposure, and then declined. No details on levels of ethanol in expired air were reported.

Applicant's summary and conclusion

Conclusions:

Overall there were some weak, but statistically significant, effects of ethanol on certain measures of neurobehavioural performance (including learning and memory, psychomotor skills and attention) in volunteers at blood alcohol concentrations of about 0.5 g/L or lower. However, no statistically significant effects were observed in rats at the same blood level (achieved in those rats receiving the lowest dose of 0.5 g/kg bw), although McKee et al. note that "although higher doses were required to produce effects in the rats, the effects observed in the rats were much more profound than those observed in humans...thus, the apparent differences between humans and rodents seem more related to the sensitivity of the test methods than to real species differences".

Executive summary:

A range of tests including standardised observational measures, spontaneous motor activity assessments, and learned visual discrimination performance was utilized in rat studies to evaluate acute CNS effects. Groups of rats were given ethanol at levels of approximately 0.5, 1.0 or 2.0 g/kg bw. In a human volunteer study, 12 healthy male subjects were given 0.65 g ethanol/kg bw and neurobehavioural effects were measured prior to and 1 and 3 hours after ethanol administration, with a computerised neurobehavioural test battery. Results of the behavioural tests in rats provided evidence of ethanol-induced changes in neuromuscular, sensori-motor, and activity domains. There were also significant changes in visual discrimination, particularly in the areas of general measures of responding and psychomotor speed. In humans, there were small but statistically significant effects on learning and memory, psychomotor skills and attention. These effects were subtle, however, and not all parameters within given domains were affected. The studies demonstrated a qualitative similarity in response to ethanol between rats and humans.