Trimethyl[2-(phosphonooxy)ethyl]ammonium chloride, calcium salt (1:1)

Basic toxicokinetics

Administrative data

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1944

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

1944

Report date:

1944

Materials and methods

Objective of study:

distribution

metabolism

Test guideline

Qualifier:

no guideline available

Principles of method if other than guideline:

The experiments were designed to follow the phosphorus from phosphorylcholine into the various blood phosphorus fractions through a period of 24 hours, to determine its distribution among representative tissues after 12 hours, and its partition between various phospholipids through several periods of time.

GLP compliance:

no

Test material

Test material information

Constituent 1

Reference substance name:

Trimethyl[2-(phosphonooxy)ethyl]ammonium chloride, calcium salt (1:1)

EC Number:

225-403-0

EC Name:

Trimethyl[2-(phosphonooxy)ethyl]ammonium chloride, calcium salt (1:1)

Cas Number:

4826-71-5

Molecular formula:

C5H13NO4P.Ca.Cl

IUPAC Name:

calcium chloride [2-(trimethylazaniumyl)ethyl] phosphate

Test material form:

solid

Specific details on test material used for the study:

Phosphorylcholine was administered as the calcium chloride salt, phosphate in the form of sodium acid phosphate (Groups II and IV) or magnesium ammonium phosphate (Group III), and phosphate plus choline as choline chloride and sodium acid phosphate or magnesium ammonium phosphate.

Radiolabelling:

yes

Test animals

Species:

rat

Strain:

not specified

Details on species / strain selection:

Albino rats

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: adult
- Weight at study initiation: see table 1
- Diet (e.g. ad libitum): standard fox chow diet ad libitum
- Water (e.g. ad libitum): ad libitum

Administration / exposure

Route of administration:

intraperitoneal

Vehicle:

other: isotonic saline

Details on exposure:

- The animals were divided into four main groups. Each group, with the exception of Group I (phosphorylcholine), was subdivided equally into three subgroups which received phosphorylcholine, phosphate, and phosphate plus choline respectiviely (see Table 1). All soluions injected were about neutral and med up in isotonic saline.

Duration and frequency of treatment / exposure:

24 hours after single administration

Doses / concentrations

Remarks:

See "Table 1. Tabulation of groups and doses".

No. of animals per sex per dose / concentration:

Group I: 12 rats
Group II: Subgroup PCh: 12 rats; subgroup P: 12 rats; subgroup P + Ch: 12 rats
Group III: Subgroup PCh: 8 rats; subgroup P: 8 rats; subgroup P + Ch: 6 rats
Group IV: Subgroup PCh: 4 rats; subgroup P: 4 rats; subgroup P + Ch: 3 rats

Details on study design:

- Dose selection rationale: all doses of phosphorylcholine were maintained as low as was experimentally feasible to avoid flooding the animal with a compound which, if normally present in the rat, probably exists in extremely minute amounts.

Details on dosing and sampling:

- The animals from Group I and the three subgroups of Group II were sacrified in groups of three after 30 minutes, 3, 12 and 24 hours by bleeding from the axillary vessels.
- The animals from Groups III and IV were divided into subgroups, injected as indicated in table I, and placed in individual metabolism cages arranged for collection of urine and feces. 12 hours after injection the animals were killed by exanguination.
- Tissues from the animals of Groups III and IV were removed immediately after sacrifice, weighed, and ground with sand.

Results and discussion

Main ADME results

Type:

excretion

Results:

12.9% of total dose (urine and faecal excretion)

Toxicokinetic / pharmacokinetic studies

Details on distribution in tissues:

If phosphorylcholine were not completely hydrolysed, and promptly, certain organs might be expected to pick up the unhydrolysed ester preferentially. The phosphorus of phosphorylcholine shows only small, if significant, differences in whole tissue distribution as compared to inorganic phosphorus in brain, testes, blood, muscle, eyes, heart, kidney, spleen, liver and intestine. Since a decreased uptake of radiophosphorus by liver phospholipid was found after 12 hours, the authors supposed phosphorylcholine has produced a decreased rate of turnover of phospholipid phosphorus or an enhanced turnover with a consequent fall 12 hours after injection. On the basis of evidence obtained on liver lecithin, cephalin, and sphingomyelin fractions at 12 hours, the authors were inclined to attribute the 12 hour differences to a decreased rate of turnover of liver phospholipid phosphorus. However, further work is required to decide this question.

Transfer into organs

Key result

Test no.:

#1

Transfer type:

blood/placenta barrier

Observation:

distinct transfer

Details on excretion:

After the administration of phosphocholine, 12.9% of the phosphorus was excreted in the urine and faeces (see table 3). Phosphorylcholine phosphorus is excreted more slowly than inorganic phosphate (17.4%).

Metabolite characterisation studies

Metabolites identified:

yes

Details on metabolites:

First, rapid hydrolysis of phosphorylcholine to give inorganic phosphate accounts for a large fraction of the administered compound.
Second, the hydrolysis of phosphorylcholine is only one of the several mechanisms by which this ester is removed from the circulating blood. From the radioactivity and phosphorus values for total acid-soluble and inorganic phosphate, it has been calculated that in the phosphorylcholine subgroup after 30 minutes only 1.3 per cent of the radiophosphorus administered was present in organic form in the isolated blood. This must indicate that a maximum of 1.3 per cent of the administered ester remained in the acid-soluble blood fraction after this period. The ester must disappear rapidly from the circulation, partially through the hydrolysis noted above as well as by diffusion on the intact ester. The metabolic utilization of the ester without liberation of inorganic phosphate would account for a further fraction.
The stepwise oxidation of phosphorylcholine through betaine aldehyde phosphate to betaine phosphate would evolve an energy-rich phosphate, which by cleavage on the proper enzyme substrate might provide the energy source for methyl transfer. Glycine, which would be formed through such a mechanism, is probably the normal demethylation product of choline.

Any other information on results incl. tables

Table 2. Specific activities and phosphorus percentages for whole tissues of rats of groups III and IV.

Tissue	Subgroup	Average P (range) (%)	Specific activity (range)	Tissue	Subgroup	Average P (range) (%)	Specific activity (range)
Group III
Liver	PCh	0.29 (0.27-0.32)	457 (354-570)	Muscle	PCh	0.28 (0.25-0.32)	133 (104-150)
	P	0.48 (0.45-0.51)	266 (233-302)		P	0.26 (0.22-0.31)	140 (96-190)
	P + Ch	0.37 (0.31-0.37)	460 (421-490)		P + Ch	0.24 (0.23-0.28)	179 (155-226)
Intestine	PCh	0.28 (0.26-0.33)	354 (263-400)	Heart	PCh	0.24 (0.22-0.27)	314 (186-403)
	P	0.29 (0.25-0.34)	416 (355-488)		P	0.22 (0.19-0.27)	295 (267-328)
	P + Ch	0.31 (0.30-0.32)	452 (439-466)		P + Ch	0.21 (0.20-0.22)	399 (391-411)
Kidney	PCh	0.32 (0.28-0.34)	319 (231-402)	Brain	PCh	0.31 (0.27-0.35)	30.2 (20.2-36.6)
	P	0.30 (0.27-0.34)	270 (223-321)		P	0.33 (0.32-0.34)	24.1 (19.5-28.6)
	P + Ch	0.31 (0.28-0.33)	356 (333-379)		P + Ch	0.32 (0.30-0.33)	28.6 (25.3-32.5)
Spleen	PCh	0.38 (0.31-0.42)	282 (175-458)	Blood	PCh	0.046 (0.041-0.050)	520 (447-580)
	P	0.35 (0.29-0.40)	301 (227-382)		P	0.047 (0.029-0.040)	438 (332-527)
	P + Ch	0.34 (0.31-0.37)	449 (386-513)		P + Ch	0.045 (0.039-0.050)	574 (548-590)
Group IV
Testes	PCh	0.22 (0.21-0.25)	135 (114-159)	Eyes	PCh	0.12	126
	P	0.22 (0.21-0.23)	104 (93-115)		P	0.12	136
	P + Ch	0.22 (0.20-0.24)	108 (93-121)		P + Ch	0.12	138

 

Table 3. Urinary and fecal excretion of P³²12 hours after administration.

Subgroup	No. of rats	Combined urinary and fecal excretion
		Average (% of total dose)	Range (% of total dose)
PCh	12	12.9	9.1-18.2
P	12	17.4	12.5-23.7
P + Ch	9	17.7	14.4-23.2

 

Applicant's summary and conclusion

Conclusions:

Phosphorylcholine phosphorus is hydrolysed rapidly to give inorganic phosphorus in the circulating blood of the rat following intraperitoneal injection of the ester. Phosphorylcholine phosphorus is excreted more slowly than inorganic phosphate, however, the phosphorus of phosphorylcholine shows only small, if significant, differences in whole tissue distribution as compared to inorganic phosphorus in brain, testes, blood, muscle, eyes, heart, kidney, spleen, liver and intestine.

Executive summary:

The metabolism of phosphorylcholine has been studied and published. 81 albino rats were divided into four main groups. Each group, with the exception of Group I (phosphorylcholine), was subdivided equally into three subgroups, which received phosphorylcholine, phosphate, and phosphate plus choline respectiviely. All soluions injected intraperitoneally and were about neutral and med up in isotonic saline. Animals from groups I and II were designed for blood experiments, and groups III and IV for distribution and excretion studies. First, a large amount of the administered phosphorylcholine hydrolyses rapidly to give inorganic phosphate. The hydrolysis of the compound is only one of the several mechanisms by which this ester is removed from the circulating blood. The ester disappears rapidly from the circulation, partially through the hydrolysis as well as by diffusion of the intact ester. Phosphorylcholine phosphorus is excreted in urine and feces more slowly than inorganic phosphate (12.9% vs. 17.4%). However, the phosphorus of phosphorylcholine shows only small, if significant, differences in whole tissue distribution as compared to inorganic phosphorus in brain, testes, blood, muscle, eyes, heart, kidney, spleen, liver and intestine.