Gum turpentine oil, consisting of the volatile fraction resulting from the distillation of oleoresin collected by tapping softwoods from pinacea derivatives (genus: pinus). It consists of terpenes, mainly monoterpenes such as alpha-pinene and beta-pinene.

Endpoint summary

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

1996

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Well described and well conducted in vivo study performed in humans, giving valuable data about absorption and excretion of several terpenes after inhalation exposure.

Reason / purpose for cross-reference:

reference to same study

Reason / purpose for cross-reference:

reference to other study

Objective of study:

absorption

excretion

Principles of method if other than guideline:

Absorption and excretion rates of alpha-pinene, beta-pinene and 3-carene after inhalation exposure to turpentine vapour in human volunteers.

GLP compliance:

no

Radiolabelling:

no

Species:

human

Sex:

male

Details on test animals or test system and environmental conditions:

Eight healthy men, not occupationally exposed to solvents, with a mean (range) age of 28 (18-37) years and weight of 75 (62-83) kg participated in the study. All were nonsmokers and they were instructed to refrain from alcohol and activities that could cause exposure to organic solvents during the preceding two days and throughout the experiment.

Route of administration:

inhalation: vapour

Vehicle:

unchanged (no vehicle)

Details on exposure:

The subjects were exposed to turpentine for two hours (450 mg/m3 (75 ppm)) during light physical exercise (50 W on a bicycle ergometer). The
exposure was conducted in an exposure chamber. The volume of the chamber was 12 m3. The air was changed 10 times/h. To prevent the solvent from leaking from the chamber into the surroundings the inlet flow was 115 m3/h and the outlet flow was 135 m3/h. The solvent was injected into the inflowing air stream by means of a high-performance liquid chromatography pump (Gilson 302). The mean temperature in the chamber was 21°C and varied by less than 2.5°C during a two hour exposure to turpentine and between different days.

Duration and frequency of treatment / exposure:

2 h, once

Remarks:

Doses / Concentrations:
450 mg/m3 (75 ppm)

No. of animals per sex per dose / concentration:

8

Control animals:

no

Positive control reference chemical:

None

Details on study design:

A low concentration of 3-carene, 10 mg/m3, was used as a control. The control concentration was chosen as low as possible so that no effect was to be expected, but the subjects could still smell the solvent.
The workload 50 W was chosen to imitate the physical activity generally found in light industries.

Details on dosing and sampling:

The solvent concentration in the air of the chamber was continuously monitored at atmospheric pressure by an infrared spectrophotometer (Miran 80) equipped with a 20 m thermostatically controlled (45°C) gas cell. Measurements of monoterpenes in the air of the chamber were made at 3.39 µm with 3.6 µm as the reference wavelength. The concentration of monoterpenes varied by less than 2% during the two hour exposure of turpentine and less than 3% between different days.

Air sampling: at four regular time intervals during the exposure exhaled air was individually collected in polyester laminated aluminium foil bags (about 20 L) for about four minutes each. Exhaled air samples were also collected after exposure (5, 15, 25, 85, 150, and 230 minutes after the end of exposure). The volume of the expired air was measured with a balanced spirometer and the concentrations of monoterpenes were analysed on a gas chromatograph (Carlo Erba, Fractovap 2350) equipped with a flame ionisation detector and a polar column (Supelcowax 10, 30 m, 0-75 mm inner diameter).

Blood sampling: arterial capillary blood (200 µL) was collected from a prewarmed fingertip at selected intervals during the exposure and up to 21 h after end of exposure, for measurement of monoterpenes in blood. Blood sampling during exposure was taken by the subject holding out one hand through a closable hole in the wall. Blood samples were collected in heparinised capillary tubes (2 x 100 µL) and transferred to gas tight head space vials (22-4 mL) which were capped immediately with Teflon lined membranes. The samples were kept for 20 minutes at 37°C before analysis.
The headspace of the blood samples was analysed on a gas chromatograph (Perkin Elmer 8700) equipped with a head space autosampler (Perkin Elmer HS-101), a polar column (Supelcowax 10, 30 m, 0-75 mm inner diameter), and a flame ionisation detector. The concentration of monoterpenes in the blood was measured by comparison with individual standard curves prepared in the same concentration range by adding monoterpenes to blood samples taken before the exposure.

Statistics:

None

Preliminary studies:

None

Details on absorption:

The relative net uptakes of the total inhaled alpha-pinene, beta-pinene, and 3-carene averaged 62%, 66%, and 68% respectively (% of total inhaled).

Details on distribution in tissues:

None

Details on excretion:

Between 2% and 8% of the net uptake was excreted unchanged in the expired air after the end of exposure. The mean blood clearance 21 h after exposure (CL21h) of alpha-pinene, beta-pinene and 3-carene, were 0.8, 0.5, and 0.4 L/kg/h, respectively. The mean half lives (t1/2) of the last phase of alpha-pinene, beta-pinene, and 3-carene averaged 32, 25, and 42 h, respectively.

Metabolites identified:

not measured

Details on metabolites:

None

The mean total concentration of the monoterpenes in the air of the chamber from all the turpentine exposures was 449 mg/m3, with a relative SD of 1-4%, based on air concentrations from four different occasions at each exposure.

If the assumption is made that the turpentine consists of alpha-pinene, beta-pinene, and 3-carene, the proportions of the vapour were about 54%, 11%, and 35%, respectively.

Table 1: Mean (SD) experimental results ofsome physiological and toxicokinetic vaiables from two hour inhalation exposure to 450 mg/m3 of turpentine during physical exercise at a workload of 50 W (also, results from previous single exposure to alpha-pinene and 3-carene are presented)

	Alpha-pinene			Beta-pinene   Exposure to turpentine	3-carene
	Exposure to turpentine	Exposure to alpha-pinene	Exposure to alpha-pinene		Exposure to turpentine	Exposure to 3-carene	Exposure to 3-carene
Air concentration (mg/m3)	242 (3.3)	455 (5.3)	225 (5.1)	49 (0.67)	157 (2.1)	451 (5.8)	228 (5.8)
Net uptake (% of total inhaled)	62 (5.3)	58 (5.3)	60 (3.9)	66 (5.8)	68 (7.5)	71 (4.7)	70 (4.4)
Respiratory elimination after exposure (% of net uptake)	3.8 (1.1)	7.7 (3.1)	5.7 (5.7)	5.0 (5.8)	2.4 (1.5) *	4.8 (1.2)	1.9 (1.5)
Concentration at steady state (30 min before end of exposure) (µmol/L)	9.0 (0.75)	19 (3.6)	9.6 (1.9)	2.6 (0.29)	9.9 (0.89)	25 (0.92)	12 (1.4)

* P<0.05, Student's t test v. pure 3-carene (450 mg/m3)

Table 2: mean (SD) apparent CLs and t1/2s of alpha-pinene, beta-pinene, and 3-carene from two hour inhalation exposure to 450 mg/m3 of alpha-pinene, 3-carene or turpentine during physical exercise at a workload of 50 W

	Alpha-pinene		Beta-pinene   Exposure to turpentine	3-carene
	Exposure to turpentine	Exposure to alpha-pinene		Exposure to turpentine	Exposure to 3-carene
CL4h (L/kg/h)	1.6 (0.3)	1.4 (0.31)	0.8 (0.3)	1.0 (0.2)	0.3 (0.1)
CL21h (L/kg/h)	0.8 (0.09) §	1.1 (0.2)	0.5 (0.3) §	0.4 (0.1) §	0.9 (0.3)
t1/2 terminal (h)	32(18)	12 (4)	25 (18)	42 (23)	30 (22)

§ four subjects due to analytical problems resulting in problems calculating AUC

Conclusions:

The mean relative uptakes of alpha-pinene and beta-pinene were 62% and 66% respectively, of the amount inhaled. Between 2% and 5 % of the net uptake was excreted unchanged in the expired air after the end of exposure. The mean blood clearance 21 h after exposure (CL21h) of alpha-pinene and beta-pinene were 0.8 and 0.5 L/kg/h, respectively. The mean half lives (t1/2) of the last phase of alpha-pinene and beta-pinene averaged 32 and 25 h, respectively.

Executive summary:

In this study, eight male volunteers were exposed to 450 mg/m3 turpentine by inhalation (2 h with physical exercise workload of 50 W) in an exposure chamber and the extent of alpha-pinene and beta-pinene inhaled as well as their excretion rates from blood were investigated.

The mean relative uptakes of alpha-pinene and beta-pinene were 62% and 66% respectively, of the amount inhaled. Between 2% and 5 % of the net uptake was excreted unchanged in the expired air after the end of exposure.

The mean blood clearance 21 h after exposure (CL21h) of alpha-pinene and beta-pinene were 0.8 and 0.5 L/kg/h, respectively. The mean half lives (t1/2) of the last phase of alpha-pinene and beta-pinene averaged 32 and 25 h, respectively.

The last phase of the t1/2s tended to be longer after exposure to turpentine than monoterpenes.

Reference 2

Endpoint:

dermal absorption in vitro / ex vivo

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Justification for type of information:

Beta-Pinene is a main constituent of UVCB substance gum tupentine oil. Therefore, data on beta-pinene can be used for extrapolation to gum turpentine oil. See read-across justification document in section 13.

Reason / purpose for cross-reference:

read-across source

Signs and symptoms of toxicity:

not specified

Dermal irritation:

not specified

Absorption in different matrices:

The dermal penetration of pure terpenes was studied during 4 h. The terpenes were present on the skin in infinite doses and the system was protected against evaporation. During that time no terpenes were detected in the acceptor fluid but extensive accumulation in the skin tissue occurred (see table 1).
Analysis of stratum corneum (SC) collected with an adhesive tape and merged into three groups demonstrates cumulation of terpenes in the outer (SC I), middle (SC II) and inner (SC III) layers. The results demonstrate rapid penetration of terpenes not only to the SC I layers but also to viable epidermis and dermis. The distance-dependent decreasing gradient of concentration for all terpenes is observed, although the concentrations were not normalized in respect of the collected SC mass. A steady-state concentration of terpenes in the SC can be assumed as soon as after 1 h. Maximum concentration in the SC was achieved as soon as after 1 h and did not further increase in the course of the study.
All studied terpenes are absorbed in high amounts in the viable epidermis with dermis (ED), however penetration into this layers is time-dependent process, constantly increasing during 4 h.

Total recovery:

No data

Conversion factor human vs. animal skin:

None

Table 1: Absorption of beta-pinene (mg/cm2) into human skin layers (mean ± S.D., n = 4)

Skin layer	1-h exposure	2-h exposure	4-h exposure
SC I	19.4 ± 5.6	28.8 ± 14.3	23.8 ± 15.7
SC II	10.5 ± 2.1	20.3 ± 11.0	12.2 ± 7.0
SC III	9.9 ± 5.4	23.5 ± 2.2	10.5 ± 3.0
SC total	39.8 ± 8.8	72.6 ± 12.1	46.5 ± 25.5
ED	89.0 ± 12.4	318.2 ± 103.4	417.8 ± 28.8
Skin total	128.8 ± 4.5	390.8 ± 106.7	464.3 ± 46.7

Table 2: Elimination of beta-pinene (mg/cm2) from human skin layers following 1 h absorption (t = 0) (mean ± S.D., n = 4)

	Time after 1-h exposure
Skin layer	0	1	2	3	4
SC I	19.4 ± 5.6	14.9 ± 6.2	12.0 ± 2.2	11.1 ± 4.2	4.8 ± 4.8
SC II	10.5 ± 2.1	6.6 ± 6.0	5.8 ± 5.0	5.5 ± 6.5	0
SC III	9.9 ± 5.4	5.1 ± 4.9	0	0	0
SC total	39.8 ± 8.8	26.6 ± 16.3	17.8 ± 7.1	16.6 ± 10.3	4.8 ± 4.8
ED	89.0 ± 12.4	84.0 ± 9.8	83.8 ± 7.4	78.1 ± 5.1	64.5 ± 4.5
Skin total	128.8 ± 4.5	110.6 ± 21.0	101.6 ± 1.7	94.7 ± 9.7	69.3 ± 8.0

Conclusions:

beta-Pinene absorption into the different skin layers is rapid (steady-state concentrations in the skin obtained after 1-h exposure) but do not permeate through the skin to the acceptor medium due to large accumulation into the skin tissue.

Executive summary:

Skin absorption and elimination kinetics were studied using human skin from the region of thorax of 40-50-years old Caucasian women, mounted on flow-through Teflon diffusion cells. beta-Pinene (500 mg) was applied onto the human skin (0.65 cm²), and after 1 to 4-h exposure, the content in the stratum corneum layers (separated by a tape-stripping method) and in the epidermis/dermis was determined using GC. Similarly, the elimination kinetics in the skin were analysed during 4 h following 1 h absorption. Quadruplicates were used for each time point.

The results demonstrate rapid penetration of terpenes not only to the first stratum corneum layers but also to viable epidermis and dermis (steady-state concentrations assumed to be obtained at 1-h exposure). However, beta-pinene did not permeate across the skin to the acceptor medium due to large cumulation in the skin tissue. Two mechanisms of elimination process of terpenes from the SC are suggested: evaporation and slightly progressive penetration from inner layer into dermis.

Reference 3

Endpoint:

dermal absorption in vitro / ex vivo

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Justification for type of information:

Alpha-Pinene is a main constituent of UVCB substance gum tupentine oil. Therefore, data on alpha-pinene can be used for extrapolation to gum turpentine oil. See read-across justification document in section 13.

Reason / purpose for cross-reference:

read-across source

Signs and symptoms of toxicity:

not specified

Dermal irritation:

not specified

Absorption in different matrices:

The dermal penetration of pure terpenes was studied during 4 h. The terpenes were present on the skin in infinite doses and the system was protected against evaporation. During that time no terpenes were detected in the acceptor fluid but extensive accumulation in the skin tissue occurred (see table 1).
Analysis of stratum corneum (SC) collected with an adhesive tape and merged into three groups demonstrates cumulation of terpenes in the outer (SC I), middle (SC II) and inner (SC III) layers. The results demonstrate rapid penetration of terpenes not only to the SC I layers but also to viable epidermis and dermis. The distance-dependent decreasing gradient of concentration for all terpenes is observed, although the concentrations were not normalized in respect of the collected SC mass. A steady-state concentration of terpenes in the SC can be assumed as soon as after 1 h. Maximum concentration in the SC was achieved as soon as after 1 h and did not further increase in the course of the study.
All studied terpenes are absorbed in high amounts in the viable epidermis with dermis (ED), however penetration into this layers is time-dependent process, constantly increasing during 4 h.

Total recovery:

No data

Conversion factor human vs. animal skin:

None

Table 1: Absorption of alpha-pinene (mg/cm2) into human skin layers (mean ± S.D., n = 4)

Skin layer	1-h exposure	2-h exposure	4-h exposure
SC I	4.3 ± 0.8	3.4 ± 0.4	7.7 ± 4.3
SC II	4.0 ± 0.5	3.2 ± 0.6	6.5 ± 4.1
SC III	3.0 ± 0.4	3.4 ± 0.8	6.0 ± 4.0
SC total	11.3 ± 4.7	10.0 ± 2.1	18.2 ± 12.1
ED	66.4 ± 16.8	147.2 ± 25.9	313.7 ± 38.3
Skin total	77.7 ± 14.5	157.2 ± 25.8	331.9 ± 34.7

Table 2: Elimination of alpha-pinene (mg/cm2) from human skin layers following 1 h absorption (t = 0) (mean ± S.D., n = 4)

	Time after 1-h exposure
Skin layer	0	1	2	3	4
SC I	4.3 ± 0.8	3.2 ± 1.9	2.0 ± 1.8	0.7 ± 0.6	0.6 ± 0.7
SC II	4.0 ± 0.5	0.9 ± 1.0	0	0	0
SC III	3.0 ± 0.4	0.2 ± 0.2	0	0	0
SC total	11.3 ± 4.7	4.3 ± 3.0	2.0 ± 1.8	0.7 ± 0.6	0.6 ± 0.7
ED	66.4 ± 16.8	45.7 ± 13.0	35.9 ± 8.2	37.3 ± 3.6	33.9 ± 5.7
Skin total	77.7 ± 14.5	50.0 ± 10.1	37.9 ± 6.8	38.0 ± 3.3	34.5 ± 5.1

Conclusions:

alpha-Pinene absorption into the different skin layers is rapid (steady-state concentrations in the skin obtained after 1-h exposure) but do not permeate through the skin to the acceptor medium due to large accumulation into the skin tissue.

Executive summary:

Skin absorption and elimination kinetics were studied using human skin from the region of thorax of 40-50-years old Caucasian women, mounted on flow-through Teflon diffusion cells. alpha-Pinene (500 mg) was applied onto the human skin (0.65 cm²), and after 1 to 4-h exposure, the content in the stratum corneum layers (separated by a tape-stripping method) and in the epidermis/dermis was determined using GC. Similarly, the elimination kinetics in the skin were analysed during 4 h following 1 h absorption. Quadruplicates were used for each time point.

The results demonstrate rapid penetration of terpenes not only to the first stratum corneum layers but also to viable epidermis and dermis (steady-state concentrations assumed to be obtained at 1-h exposure). However, alpha-pinene did not permeate across the skin to the acceptor medium due to large cumulation in the skin tissue. Two mechanisms of elimination process of terpenes from the SC are suggested: evaporation and slightly progressive penetration from inner layer into dermis.

Reference 4

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

1981

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Results from publication with well described details

Reason / purpose for cross-reference:

reference to same study

Reason / purpose for cross-reference:

reference to other study

Objective of study:

toxicokinetics

Principles of method if other than guideline:

The blood and urine monoterpene concentrations were continuously monitored from a patient attempting suicide by ingestion of 400-500 mL pine oil.

GLP compliance:

no

Radiolabelling:

no

Species:

human

Sex:

male

Details on test animals or test system and environmental conditions:

A 49 year old male (height 185 cm, body weight 85 kg)

Route of administration:

oral: unspecified

Vehicle:

unchanged (no vehicle)

Duration and frequency of treatment / exposure:

Once

Remarks:

Doses / Concentrations:
400-500 mL of pine oil

No. of animals per sex per dose / concentration:

1

Control animals:

no

Preliminary studies:

None

Details on absorption:

See other information on results

Details on distribution in tissues:

See other information on results

Details on excretion:

See other information on results

Metabolites identified:

yes

Details on metabolites:

See other information on results

Clinical signs: psychomotric excitation, headache, erythem of mouth and larynx, a flush of the face, ataxia, and a spontaneous hyperventilation. With a latence of 10 h after ingestion the consciousness of the patient was impaired and the circulatory parameters became instable although a hypovolemy could be excluded. Three weeks later the patient left the clinic without any bodily complaints.

Examination results: the circulatory parameters and the laboratory data were in the normal range. The EEG recorded the second day revealed a decelerated activity. No epileptogenic activities could be detected. The patient had a retrograde amnesia for the period of somnolence and sopor. At this time a leukocytosis (21000/mm3), a slight raise of the transaminases, and a reduction of the pseudocholinesterase (1446 U/L) were observed. The renal functions were not affected except a transient oliguria which was due to the drop of the blood pressure.

Alpha-pinene was the monoterpene with the highest concentration in blood. The high affinity of the monoterpenes to lipophilic body compartments may be partly responsible for the comparatively low alpha-pinene blood level. The metabolites of the monoterpenes could be detected solely in urine whereas in blood only the original monoterpenes were identified. The main metabolite is bornylacetate. Ten days after ingestion a total of 100 mg bornylacetate was excreted with the urine. The renal excretion of metabolized monoterpenes reaches its peak level 5 days after ingestion. This indicates that the resorbed portion of the monoterpenes is slowly metabolized and then excreted via kidneys. The main metabolic pathways are hydratation, hydroxylation, rearrangement, and acetylation.

Conclusions:

The data suggest that monoterpenes are poorly absorbed in the gastrointestinal tract. The absorbed portion of the hydrocarbons accumulates in the lipophilic body compartments and is slowly metabolized and then excreted by the kidneys. The main metabolic pathways are hydratation, hydroxylation, rearrangement, and acetylation. Five metabolites were identified.

Executive summary:

A patient attempting suicide ingested 400-500 mL pine oil and was admitted to the clinic. Since more than the potentially lethal dose had been ingested hemoperfusions with activated charcoal and amberlite and a hemodialysis were performed. Clinical signs observed were: psychomotric excitation, head ache, erythem of mouth and larynx, a flush of the face, ataxia, and a spontaneous hyperventilation. With a latence of 10 h after ingestion the consciousness of the patient was impaired and the circulatory parameters became instable although a hypovolemy could be excluded. Three weeks later the patient left the clinic without any bodily complaints. The composition of the ingested pine oil was determined by gaschromatography/mass spectrometry. Four monoterpenes were identified in the pine oil ingested: 57% alpha-pinene, 8% beta-pinene, 26% carene, 6% limonene and 3% other hydrocarbons. The blood and urine monoterpene concentrations were continuously monitored. The data suggest that monoterpenes are poorly absorbed in the gastrointestinal tract. The absorbed portion of the hydrocarbons accumulates in the lipophilic body compartments and is slowly metabolized and then excreted by the kidneys. The main metabolic pathways are hydratation, hydroxylation, rearrangement, and acetylation. Five metabolites were identified.

Reference 5

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

1967

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Old but well conducted and well described study.

Reason / purpose for cross-reference:

reference to same study

Reason / purpose for cross-reference:

reference to other study

Objective of study:

distribution

Principles of method if other than guideline:

Turpentine content of tissues after chronic exposure of adult male rats to commercial turpentine.

GLP compliance:

no

Radiolabelling:

no

Species:

rat

Strain:

Wistar

Sex:

male

Details on test animals or test system and environmental conditions:

No food was offered during the exposures while water was given ad libitum. The animals were kept under an artificial day-light cycle of 12 h for 2 weeks before and during the exposures.

Route of administration:

inhalation: vapour

Vehicle:

unchanged (no vehicle)

Duration and frequency of treatment / exposure:

6 h /day, 5 days/ weeks for 8 weeks

Remarks:

Doses / Concentrations:
300 ppm (12 µmol/L)

No. of animals per sex per dose / concentration:

40

Control animals:

yes, concurrent no treatment

Positive control reference chemical:

None

Details on dosing and sampling:

5 rats were killed by decapitation 1, 4, 5, 6, 7 and 8 weeks after the beginning of the experiment while ten rats were killed at the second week. Brain and perinephric fat samples were taken after killing.
The amount of alpha-pinene was analyzed gas-chromatographically after extraction with dimethylformamide (DMFA).

Statistics:

None

Preliminary studies:

None

Details on absorption:

None

Details on distribution in tissues:

Alpha-pinene was found in the perinephric fat and brain. The brain pinene content remained similar throughout the experiment and it was about 10% of that in fat.

Details on excretion:

None

Metabolites identified:

not measured

Details on metabolites:

None

Table 1: mean alpha-pinene content in brain and perinephric fat

Weeks of exposure	Alpha-pinene in brain (mmol/g)	Alpha-pinene in perinephric fat (mmol/g)
1	26±6	365±85
2	28±10	353±110
4	27±7	307±76
5	21±4	322±72
6	23±4	371±65
7	-	-
8	-	-

Conclusions:

Chronic exposure of adult male rats to commercial turpentine resulted in an accumulation of the solvent in perinephric fat and brain.

Executive summary:

Male wistar rats were exposed to 300 ppm of turpentine by inhalation 6 h/day, 5days/week for 8 weeks. 5 rats were killed by decapitation 1, 4, 5, 6, 7 and 8 weeks after the beginning of the experiment while ten rats were killed at the second week. Brain and perinephric fat samples were taken after killing. The amount of alpha-pinene was analyzed gas-chromatographically after extraction with dimethylformamide (DMFA). After chronic exposure of adult male rats, commercial turpentine was found in perinephric fat and brain. The brain pinene content remained similar throughout the experiment and it was about 10% of that in fat.

Reference 6

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

1967

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Well conducted study but with few details on test material.

Reason / purpose for cross-reference:

reference to same study

Reason / purpose for cross-reference:

reference to other study

Objective of study:

distribution

Principles of method if other than guideline:

Turpentine concentrations in rat tissues were determined immediately after 1 and 2 hours of exposure, and after 15, 30, and 60 minutes post-exposure using at least three animals for each of these periods.

GLP compliance:

no

Radiolabelling:

no

Species:

rat

Strain:

Wistar

Sex:

male

Details on test animals or test system and environmental conditions:

Male Wistar rats 140-200 g were exposed to flow rates of about 150-200 mL/min

Route of administration:

inhalation: vapour

Vehicle:

unchanged (no vehicle)

Duration and frequency of treatment / exposure:

1 or 2 h, once

Remarks:

Doses / Concentrations:
1-h exposure: 10-12 mg/L
2-h exposure: 11-13 mg/L

No. of animals per sex per dose / concentration:

3

Control animals:

yes

Positive control reference chemical:

None

Statistics:

None

Preliminary studies:

None

Details on absorption:

None

Details on distribution in tissues:

In rats, the highest concentrations of inhaled turpentine were found in the spleen, brain, kidneys and liver. The low blood level might indicate rapid tissue uptake and slow release from tissues.

Details on excretion:

Lung concentrations were higher after 30 min than after 15 min which might indicate that lung excretion was occuring. The low blood level might indicate rapid tissue uptake and slow release from tissues.

Metabolites identified:

not measured

Details on metabolites:

None

Table 1: concentration of turpentine in tissues of rats at various intervals fter 1 or 2-h exposure to turpentine vapour

		Concentration after 1-hour exposure (µg/g)				Concentration after 2-hour exposure (µg/g)
Tissue	Mean recovery (%) *	0	15 min	30 min	60 min	0	15 min	30 min	60 min
Brain	25	160	63	49	20	127	47	21	15
Spleen	50	214	127	39	19	94	32	15	17
Kidney	35	146	58	26	0	97	26	8	12
Liver	15	167	43	33	0	157	34	17	8
Lung	50	101	0	26	0	54	20	25	7
Blood	65	24	16	8	1	4	0.4	0.7	0.9

* refers to recovery of turpentine added to the corresponding tissues of unexposed animals. The values given for the test animals are corrected for this incomplete recovery.

Conclusions:

In rats, the highest concentrations of inhaled turpentine were found in the spleen, brain, kidneys and liver. Lung concentrations were higher after 30 min than after 15 min which might indicate that lung excretion was occuring. The low blood level might indicate rapid tissue uptake and slow release from tissues.

Executive summary:

Turpentine concentrations in rat tissues were determined immediately after 1 and 2 hours of exposure to 10 to 13 mg/L of turpentine vapour, and after 15, 30, and 60 minutes post-exposure using at least three animals for each of these periods.

In rats, the highest concentrations of inhaled turpentine were found in the spleen, brain, kidneys and liver. Lung concentrations were higher after 30 min than after 15 min which might indicate that lung excretion was occuring. The low blood level might indicate rapid tissue uptake and slow release from tissues.

Description of key information

Short description of key information on bioaccumulation potential result:
Turpentine oil, alpha-pinene and beta-pinene are readily absorbed by inhalation.

Only a minor part is exhaled unchanged and even less is excreted unchanged in urine, showing that they are extensively metabolised. However, they all show high affinity to adipose tissues which entails a rapid distribution in fat tissues and a long terminal half-life from blood because of a slow release from fat tissues.

Short description of key information on absorption rate:
Results demonstrate rapid penetration of pinenes not only to the first stratum corneum layers but also to viable epidermis and dermis. In an in vitro study, alpha and beta-pinene did not permeate across the skin to the acceptor medium due to large cumulation in the skin tissue. However, following immersion of young pigs and one human subject for 30 minutes in baths containing 150 mL of a pine-oil mixture, alpha- and beta-pinene were detected in exhaled air within 20 minutes reaching maximum levels 50-75 minutes after start of the bath and remained detectable after 1 day.

Key value for chemical safety assessment

Bioaccumulation potential:

no bioaccumulation potential

Additional information

Eight male volunteers were exposed to 450 mg/m3 turpentine by inhalation (2 h, 50 W) in an exposure chamber. The mean relative uptakes of alpha-pinene and beta-pinene were 62% and 66% respectively, of the amount inhaled. Between 2% and 5% of the net uptake was excreted unchanged in the expired air after the end of exposure. The mean blood clearance 21 h after exposure (CL21h) of alpha-pinene and beta-pinene were 0.8 and 0.5 L/kg/h, respectively. The mean half lives (t1/2) of the last phase of alpha-pinene and beta-pinene averaged 32 and 25 h, respectively (Filipsson et al., 1996).

Since turpentine is a lipophilic substance, it accumulates in fatty tissues. In rats, the highest concentrations of inhaled turpentine were found in the spleen, kidneys, brain, and perinephric fat (Sperling et al., 1967; Savolainen et al., 1978).

Elimination of turpentine and its metabolites is primarily through the urinary tract (Lewander and Aleguas, 1998, cited in HSDB 2003). A portion of the turpentine absorbed in industrial exposures is exhaled unchanged in expired air. The remainder is excreted in the urine as glucuronic acid conjugates (Bingham, 2001, cited in HSDB 2003).

The blood and urine monoterpene concentrations were continuously monitored from a patient attempting suicide by ingestion of 400-500 mL pine oil. The blood and urine monoterpene concentrations were continuously monitored. The data suggest that monoterpenes are poorly resorbed in the gastrointestinal tract. The resorbed portion of the hydrocarbons cumulates in the lipophilic body compartments and is slowly metabolized and then excreted by the kidneys. The main metabolic pathways are hydratation, hydroxylation, rearrangement, and acetylation (Koppel, 1981).

The toxicokinetics of alpha-pinene was studied in humans. Eight healthy males were exposed to 10, 225, or 450 mg/m3 (+)-alpha-pinene or 450 mg/m3 (-)-alpha-pinene for 2 h in an inhalation chamber while performing light work (50 watts). Average pulmonary uptake of (+)-alpha-pinene and (-)-alpha-pinene amounted to 59% of the exposure concentration. Absolute uptake increased linearly with concentration. Mean blood concentration at the end of exposure were linearly related to inhaled concentration. The terminal t1/2 of alpha-pinene from the blood 695 min for (+)-alpha-pinene and 555 min for (-)-alpha-pinene. Cumulative urinary excretion of unchanged alpha-pinene amounted to less than 0.001% of each dose. Respiratory elimination of (+)-alpha-pinene and (-)-alpha-pinene was 7.7 and 7.5% of total uptake, respectively (Falk et al., 1990; cited in HSDB 2003 and 2009a).

Similarly, the renal elimination of verbenols after experimental exposure to (+) and (-)alpha-pinene was studied in humans following exposure to 10, 225, and 450 mg/m3 terpene in an exposure chamber. The pulmonary uptake was about 60%. About 8% was eliminated unchanged in exhaled air. Depending on the exposure level, about 1%-4% of the total uptake was eliminated as cis- and trans-verbenol. Most of the verbenols were eliminated within 20 h after a 2-h exposure. The renal excretion of unchanged alpha-pinene was less than 0.001%. (Levin et al., 1992, cited in HSDB 2009a)

Both alpha- and beta-pinene are readily absorbed through the pulmonary system, the skin, and the intestines (Clayton and Clayton,1981-1982; Budavari et al., 1996; cited in HSDB, 2009a and 2009b)

The biotransformation of (+)-, (-)-, and (+/-)-alpha-pinenes, (-)-beta-pinene in rabbits was investigated. The major metabolites were as follows: (-)-trans-verbenol from (+)-, (-)-, and (+/-)-alpha-pinenes; (-)-10-pinanol and (-)-1-p-menthene-7,8-diol from (-)-beta-pinene (Ishida, 1981).

Dermal absorption: Results demonstrate rapid penetration of pinenes not only to the first stratum corneum layers but also to viable epidermis and dermis. In an vitro study, alpha and beta-pinene did not permeate across the skin to the acceptor medium due to large cumulation in the skin tissue (Cal et al., 2006). However, following immersion of young pigs and one human subject for 30 minutes in baths containing 150 mL of a pine-oil mixture, alpha and beta-pinene were detected in exhaled air within 20 minutes reaching maximum levels 50-75 minutes after start of the bath and remained detectable after 1 day. (Opdyke, 1979, cited in HSDB 2009)

In conclusion, bioaccumulation of these terpene hydrocarbons does not occur, since the substances are efficiently metabolised to yield oxygenated metabolites (verbenol, myrtenol and myrtenic acid) that are subsequently conjugated with glucuronic acid and excreted mainly in the urine.

HSDB (Hazardous Substances Data Bank). 2003. Turpentine. HSDB No. 204. Produced by the National Library of Medicine (NLM), Bethesda, M.D. Last updated 15 October 2003.

HSDB (Hazardous Substances Data Bank). 2009a. Alpha pinene. HSDB No. 720. Produced by the National Library of Medicine (NLM), Bethesda, M.D. Last Revision Date: 26 June 2009

HSDB (Hazardous Substances Data Bank). 2009b. beta pinene. HSDB No. 5615. Produced by the National Library of Medicine (NLM), Bethesda, M.D. Last Revision Date: 26 June 2009

Discussion on absorption rate:

Skin absorption of alpha and beta-pinene were studied using human skin mounted on flow-through diffusion cells. Similarly, the elimination kinetics in the skin were analysed during 4 h following 1-h exposure. The results demonstrate rapid penetration of pinenes not only to the first stratum corneum layers but also to viable epidermis and dermis (steady-state concentrations assumed to be obtained at 1-h exposure). However, pinenes did not permeate across the skin to the acceptor medium due to large cumulation in the skin tissue. Two mechanisms of elimination process of terpenes from the SC are suggested: evaporation and slightly progressive penetration from inner layer into dermis. (Cal et al., 2006)

Following immersion of young pigs and one human subject for 30 minutes in baths containing 150 mL of a pine-oil mixture (Fichtennadel-Latschenkieferol Kneipp) in 450 L of water, alpha-pinene, beta-pinene and limonene (components of Latschenkieferol) were detected in exhaled air within 20 minutes reaching maximum levels 50-75 minutes after start of the bath and remained detectable after 1 day. (Opdyke, 1979, cited in HSDB 2009)

HSDB (Hazardous Substances Data Bank). 2009. Alpha pinene. HSDB No. 720. Produced by the National Library of Medicine (NLM), Bethesda, M.D. Last Revision Date: 26 June 2009