(Z)-3,7-dimethylocta-2,6-dienal

Basic toxicokinetics

Administrative data

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Acceptable, well documented publication which meets basic scientific principles; study acceptable as key study

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Cross-reference 2

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

publication

Title:

Disposition of citral in male Fischer rats

Author:

Diliberto JJ, Usha G, Birnbaum LS

Year:

1988

Bibliographic source:

Drug Metab. Dispos. 16, 721-727

Materials and methods

Objective of study:

distribution

excretion

toxicokinetics

Test guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

Time course of distribution of 14C-label in tissues, blood, bile, urine, feces, expired air measured by liquid scintillation counting after single and repeated application; separation of unchanged and metabolized citral in blood and bile by HPLC (metabolites not identified)

GLP compliance:

not specified

Test material

Test material information

Constituent 1

Reference substance name:

Citral

EC Number:

226-394-6

EC Name:

Citral

Cas Number:

5392-40-5

IUPAC Name:

3,7-dimethylocta-2,6-dienal

Constituent 2

Reference substance name:

(Z)-3,7-dimethylocta-2,6-dienal

EC Number:

203-379-2

EC Name:

(Z)-3,7-dimethylocta-2,6-dienal

Cas Number:

106-26-3

Molecular formula:

C10H16O

IUPAC Name:

(2Z)-3,7-dimethylocta-2,6-dienal

Details on test material:

- Name of test material (as cited in study report): citral and 14C-citral
- Analytical purity: >= 98%, purified by HPLC and concentrated into a biologically compatible solvent using a C18 reverse phase SDep_Pak cartridge
- Impurities (identity and concentrations): no data
- Composition of test material, percentage of components:
- Isomers composition: 74% geranial, 26% neral
- Specific activity (if radiolabelling): 10.7 mCi/mmol
- Locations of the label (if radiolabelling): C1 and C2

Radiolabelling:

yes

Remarks:

(1,2-14C)-citral

Test animals

Species:

rat

Strain:

Fischer 344

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Breeding Laboratories, Portage, MI, USA
- Age at study initiation: 2-3 month
- Weight at study initiation: 200-250 g
- Fasting period before study: no data
- Housing: individually
- Individual metabolism cages: yes
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 7 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23 +/-2
- Humidity (%): 50 +/- 5
- Air changes (per hr): no data, air flow rate through the cages 0.3-0.4 L/min
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

other: oral gavage and intraveneous

Vehicle:

other: Ethanol/Emulphor EL-620/water: ratio 1:1:3 for oral application, ratio 1:1:8 for i.v. studies

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
radiolabeled citral was diluted with unlabeled citral to administer 1 µCi/rat for oral administration and 0.5 µCi/rat for i.v. administration

VEHICLE
- Concentration in vehicle: oral application 1, 10 or 100 mg/mL; i.v. application 2.5 mg/mL

Duration and frequency of treatment / exposure:

acute study: single treatment
multiple dosing study: oral pretreatment for 10 days with unlabelled citral at a dose of 5 mg/kg bw/day followed by single oral or i.v. dose of 5 mg/kg 14C-citral

Doses / concentrations

Remarks:

Doses / Concentrations:
oral application: 5, 50, 500 mg/kg/d
i.v. application: 5 mg/kg bw/d

No. of animals per sex per dose / concentration:

N ≥ 3

Details on study design:

- Dose selection rationale: 0.1, 1, and 10 % of the oral LD50 of 5000 mg/kg

Details on dosing and sampling:

PHARMACOKINETIC STUDY (distribution, excretion)
- Tissues and body fluids sampled: urine, faeces, expired air, blood, liver, kidneys, adrenals, thymus, spleen, brain, heart, lungs, testes, skin, adipose tissue, muscle, stomach contents, small intestine contents, large intestine contents, tail site (for i.v. application), bile
- Time and frequency of sampling:
excreta samples at 2, 4, 6, 8, 12, 16, 24, 32, 48, and 72 hrs;
tissue samples at sacrifice at 72 hrs p.a.
bile samples: at 5, 15, 30, 45, 60, 90, 120, 150, 180, 210, 240, 270 min after dosing by cannulation of the common bile duct
blood samples: at 1, 2, 5, 10, 15, 20, 430, 45, 60, 90, 120, 150, 180, 210 and 240 min p.a. by cannulation of the jugular vein
air samples: the total air flow through the metabolism cages was continuously passed through two consecutive traps (charcoal trap and bubbler trap, see below)
- Quantitative measurements:
tissue radioactivity: after solubilisation with TEAH (tetraethyammonium hydroxide) followed by liquid scintillation counting
urine: liquid scintillation counting
faeces: after softening with Triton-X, homogenisation and solubilisation with TEAH, followed by liquid scintillation counting
expired air: activated charcoal filter traps for expired volatiles were extracted with ethanol, followed by liquid scintillation counting;
bubbler trap solutions containing ethanolamine and ethylene glycol monomethylether (3:7) to trap expired carbon dioxide were analyzed directly by liquid scintillation counting
bile samples: total radioactivity by direct liquid scintillation counting
blood samples: after solubilisation with TEAH and concentration on a Sep-Pak cartridge the eluate was used for liquid scintillation counting
METABOLITE CHARACTERISATION STUDIES
- Tissues and body fluids sampled: bile and blood
- Time and frequency of sampling: see above
- From how many animals: 3
- Method type(s) for identification: bile samples were analyzed by HPLC to resolve into parent and metabolite fractions (no further identifcation of individual metabolites), quantitative analysis via radioactive labelling
- Limits of detection and quantification: not specified

Statistics:

computer based nonlinear regression analysis (RSI Bolt Beranek and Newman, Inc ., Cambridge, MA); additional two-tailed students t-test and z-statistics

Results and discussion

Toxicokinetic / pharmacokinetic studies

Details on absorption:

Rapid and complete absorption from gastrointestinal tract (91 - 95%). Citral was rapidly and completely absorbed after oral exposure. The difference between fecal excretion after oral and i.v. exposure indicated that 5 to 9% of the oral dose was not absorbed from the gastrointestinal tract and was eliminated via the feces.

Details on distribution in tissues:

amount in any tissue < 2%; highest concentrations in liver, muscle, blood, adipose tissue
relative amount in tissue independent of dose or route of administration

Details on excretion:

Excretion profiles independent from dose or route of administration:

Recovery after single 5 mg/kg oral dose:
24 hours: 67% with 45% in urine, 16% as exhaled 14CO2, 6% in feces, <1% as exhaled 14C-citral;
production of 14CO2 essentially ceased by 12 hrs;
72 hours: 83% (+- 10 %) with 51% in urine, 17% as exhaled 14CO2, 12% in feces, 3% in tissues, <1% as exhaled 14C-citral

Recovery after single 5 mg/kg i.v. dose:
12 hours: 57% with 47% in urine, 7% as 14CO2, 2% in feces, <1% as exhaled 14C-citral;
elimination essentially completed within 24 hrs
72 hours: 79% (+- 18 %) with 58% in urine, 8% as 14CO2, 7% via the feces, 6% tissues, <1% as exhaled 14C-citral

Elimination via bile after a single 5 mg/kg i.v. dose:
20% of the dose appeared in bile within 1 hr, with another 7% appearing by 4.5 hrs. The amount excreted in the bile was 4 times higher than that excreted in the feces within 3 days. HPLC of bile, even as early 5 min after treatment, demonstrated the complete absence of any unmetabolized citral.

Effect of multiple dosing:
In rats pretreated for 10 days (5 mg/kg bw/d orally), biliary excretion was increased to 34%, while excretion via urine, feces or expired CO2 was not affected. Therefore, repeated exposure did not alter the overall pattern of disposition.

Toxicokinetic parameters

Toxicokinetic parameters 1

Toxicokinetic parameters:

half-life 1st: 11 min for elimination from blood after i.v. application

Toxicokinetic parameters 2

Toxicokinetic parameters:

half-life 2nd: 43 min for elimination from blood after i.v. application

Toxicokinetic parameters 3

Toxicokinetic parameters:

half-life 3rd: 27 hr for elimination from blood after i.v. application

Toxicokinetic parameters 4

Toxicokinetic parameters:

half-life 1st: 1.8 h for elimination via urine after 5 mg/kg p.o.

Toxicokinetic parameters 5

Toxicokinetic parameters:

half-life 1st: 2.6 h for elimination of 14CO2 after 5 mg/kg p.o.

Toxicokinetic parameters 6

Toxicokinetic parameters:

half-life 1st: 11 h for elimination via faeces after 5 mg/kg p.o.

Metabolite characterisation studies

Metabolites identified:

not measured

Any other information on results incl. tables

Table 1: Distribution of citral-derived radioactivity in tissues and excreta after oral or i.v.exposure, 72 hr after administration (Values are means ± SE; n ≥ 3 )

Tissue or excreta	Test groups
	Oral			Preatreated^a 5 mg/kg	i.v.
	500 mg/kg	50 mg/kg	5 mg/kg		5 mg/kg
	% of administered dose
Total tissues^b	5.21 ± 1.95	2.81 ± 0.77	3.17 ± 0.99	4.44 ± 1.00	6.31 ± 0.78
Blood	0.52 ± 0.23	0.19 ± 0.05	0.36 ± 0.11	0.13 ± 0.01	0.40 ± 0.14
Liver	0.94 ± 0.25	0.58 ± 0.18	0.70 ± 0.11	0.39 ± 0.04	0.94 ± 0.07
Kidneys	0.08 ± 0.02	0.04 ± 0.01	0.07 ± 0.01	0.08 ± 0.01	0.09 ± 0.01
Skin	1.00 ± 0.61	0.46 ± 0.08	0.64 ± 0.22	0.74 ± 0.12	1.35 ± 0.01
Adipose	0.16 ± 0.09	0.24 ± 0.06	0.44 ± 0.14	0.15 ± 0.01	0.79 ± 0.12
Muscle	0.67 ± 0.04	0.63 ± 0.00	0.53 ± 0.19	1.94 ± 0.59	1.93 ± 0.56
Stomach contents	0.16  ± 0.10	< 0.01	0.06 ± 0.04	0.00	0.05 ± 0.30
Small intestine contents	0.39  ± 0.15	0.17 ± 0.08	0.03 ± 0.01	0.28 ± 0.07	0.07 ± 0.02
Large intestine contents	0.65  ± 0.11	0.13 ± 0.06	0.11 ± 0.06	0.50 ± 0.11	0.12 ± 0.02
i.v. tail site	NA^c	NA	NA	NA	0.12 ± 0.07
Urine	63.36 ± 10.23	47.92 ± 2.03	50.69 ± 5.19	52.96 ± 3.38	57.88 ± 15.02
Feces	13.22 ± 4.44	15.58 ± 2.89	11.99 ± 1.61	11.11 ± 1.56	6.84 ± 1.49
Expired CO2	12.70 ± 3.37	10.37 ± 0.72	16.62 ± 1.92	14.66 ± 0.73	7.92 ± 0.79
Expired Citral	0.16 ± 0.11	0.04 ± 0.01	0.48 ± 0.21	0.03 ± 0.01	0.33 ± 0.30
Total	93.65 ± 20.10	76.73 ± 6.42	82.95 ± 9.93	83.20 ± 6.78	79.28 ± 18.38

^a Pretreated with 5 mg of citral/kg/day po for 10 days, then po with [¹⁴C]citral.

^bTotal of collected tissues, including those listed except for dermal skin site. Tissues not individually listed contain 0.1% of the total dose.

^cNA, not applicable.

Table 2: Toxicokinetic parameters for the elimination of citral-derived radioactivity (Estimate ± SE)

Excreta	Dose [mg/kg]	Component^a	Pool size [% total dose]	Decay rate [hr^-1]	Half life [hr]
Urine	po
	5	1	47.29 ± 32.53	0.378 ± 0.124	1.8
		2	0.96 ± 0.15	0.044 ± 0.006	15.8
	50	1	23.19 ± 0.83	0.252 ± 0.006	2.8
		2	0.42 ± 0.09^b	0.029 ± 0.006	23.9
	500	1	26.94 ± 12.59	0.161 ± 0.030	4.3
		2	1.05 ± 0.52	0.015 ± 0.016	46.2
	i.v. 5	1	57.97±0.12	0.208±0.001	3.3
		2	0.64±0.16	0.046±0.008	15.1
CO2	po
	5	1	11.66 ± 0.75	0.271 ± 0.008	2.6
		2	0.14 ± 0.01	0.042 ± 0.002	16.5
	50	1	10.16 ± 0.94	0.316 ± 0.012	2.2
		2	0.09 ± 0.01^b	0.084 ± 0,004^b	8.3
	500	1	2.98 ± 0.42^b	0.138 ± 0.016^b	5.0
	i.v. 5	1	2.09±0.30	0.280±0.058	2.5
		2	0.07±0.02	0.064±0.012	13.8
Feces	po
	5	1	1.82 ± 0.89	0.063 ± 0.035	11.0
	50	1	1.11 ± 0.18	0.045 ± 0.011	15.4
	500	1	0.95 ± 0.12	0.037 ± 0.008	18.7
	i.v. 5	1	0.67±0.21	0.093±0.035	7.5

^a Component of the elimination phase: 1 = initial component of eliminiation phase; 2 = terminal component of elimination phase

^bSignificantly different from 5 mg/kg po (z test, p < 0.01)

In summary, most of the citral-derived radioactivity was rapidly eliminated from the body with a whole body half-life of 8 hr after i.v. exposure. However, a small percentage tended to persist with a clearance half-life of 24 hrs.

Elimination from blood after i.v. administration:

< 25% of administered ¹⁴C-activity remained in the blood within 2 min p.a.; within 5 min several metabolites (not identified) and no citral were found in blood; elimination from blood was characterized by a three-exponential decay curve (pool size of the first, the second and the third phase was 26+-2%, 17+-1% and 4+-1% of the total dose, respectively).