Zinc bis(dihydrogen phosphate)

Acute Toxicity: other routes

Administrative data

Endpoint:

acute toxicity: other routes

Type of information:

experimental study

Adequacy of study:

key study

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:

The biochemical and pathological changes produced by the intratracheal instillation of certain components of zinc-hexachloroethane smoke

Author:

Richards RJ, Atkins J, Marrs TC, Brown RFR & Masek L

Year:

1989

Bibliographic source:

Toxicol 54: 79-88

Materials and methods

Test guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

Biochemical and pathological study in lungs-
The potential of the test material to induce respiratory adverse effects/damage to the lungs in Porton Wistar rats was investigated using intra-tracheal instillation of the test material as a single dose. Oedema resulting from the treatment was assessed by increases in lung wet weight together with elevation in the levels of a purified alveolar surface protein fraction derived from the perfused lungs. The formation of a protein rich oedematous fluid is a common early response to toxic substances.
The toxicity of the test material can be correlated with pulmonary oedema inducing capacity, since protein rich oedematous fluid is a common early response to toxic substances.
Assessment of oedematous reaction is carried out by determination of increased in lung wet weight together with elevated levels of a purified alveolar surface protein fraction derived from lungs which had been totally perfused of blood.

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Zinc chloride

EC Number:

231-592-0

EC Name:

Zinc chloride

Cas Number:

7646-85-7

Molecular formula:

Cl2Zn

IUPAC Name:

zinc dichloride

Details on test material:

- Name of test material (as cited in study report): Zinc chloride

Test animals

Species:

rat

Strain:

Wistar

Sex:

male

Details on test animals or test system and environmental conditions:

- Weight at study initiation: 150 -200 g
- Diet (e.g. ad libitum): Commercial diet
- Water (e.g. ad libitum): Tap water

Administration / exposure

Route of administration:

other: intratracheal

Vehicle:

water

Doses:

1. Investigation of rat lung response at different dose levels: 0.25, 0.5, 1.0, 2.0, 4.0 and 5.0 mg/kg bw
2. Investigation dose levels which produce early alveolar oedema: 0.25, 0.5, 1.0 or 1.5 mg/kg bw
3. Investigation of the effects of the perfusion procedure: 1.5 mg/kg bw
4. Investigation of the time course of oedema and any signs of fibrogenesis: 2.5 mg/kg bw
5. Histopatholgical examination of lungs: 2.5 mg/kg bw

No. of animals per sex per dose:

1. Investigation of lung response at low dose: 4
2. Investigation of rat lung response at different dose levels: Not reported
3. Investigation of dose level which produces early alveolar oedema: 6
4. Investigation of the effects of the perfusion procedure: 10
5. Investigation of the time course of oedema and any signs of fibrogenesis: 6
6. Histopatholgical examination of lungs: 10

Control animals:

yes

Details on study design:

- Duration of observation period following administration:
1. To investigate lung response at low dose: 2 and 4 wk
2. To investigate rat lung response at different dose levels: 12 h and 7 d
3. To investigate dose level which produces early alveolar oedema: 20 h
4. To investigate the effects of the perfusion procedure: 20 h
5. To investigate the time course of oedema and any signs of fibrogenesis: 6, 18, 28, 48, 72 h, 7 d and 14 d
6. Histopatholgical examination of lungs: 6, 28, 72 h, 7 d and 14 d

- Necropsy of survivors performed: yes

EXAMINATION
- Examination performed for different groups:
1. To investigate lung response at low dose: Lung wet weight and alveolar surface protein levels
2. To investigate rat lung response at different dose levels: Histopathological examination
3. To investigate dose level which produces early alveolar oedema: Concentration of protein in the lavage fluid
4. To investigate the effects of the perfusion procedure: Lung wet weight and alveolar surface protein levels
5. To investigate the time course of oedema and any signs of fibrogenesis: Lung wet weight, alveolar surface protein levels after all time intervals (6, 18, 28, 48, 72 h, 7 dand 14 d) and lung dried weight and hydroxyproline in the total lavaged lung after 7 and 14 d
6. Histopatholgical examination of lungs
- Other examinations: Biochemical analysis using standard techniques

Statistics:

Statistical analysis of the data was carried out using an unpaired Student's t-test and differences between mean values were considered significant when P < 0.05.

Results and discussion

Effect levels

Effect levels 1

Sex:

male

Dose descriptor:

other: Elevated alveolar surface protein and lung wet weight

Effect level:

0.3 mg/kg bw

Remarks on result:

other: exposure duration: 14d

Effect levels 2

Sex:

male

Dose descriptor:

other: intra-alveolar oedema with charecteristic eosinophilic fluid accumulation and fibrin clots

Effect level:

> 0.5 mg/kg bw

Remarks on result:

other: exposure duration: 12h

Effect levels 3

Sex:

male

Dose descriptor:

other: interstitial fibrosis

Effect level:

2.5 mg/kg bw

Remarks on result:

other: exposure duration: 14d

Effect levels 4

Sex:

male

Dose descriptor:

other: Fibrogenic response

Effect level:

2.5 mg/kg bw

Remarks on result:

other: exposure duration: 7d

Mortality:

not reported

Clinical signs:

not observed

Body weight:

not observed

Gross pathology:

not observed

Other findings:

- Other observations:
1. Results for investigation of lung response at low dose after 2 or 4 wks: See Table 1
2. Results for investigation of rat lung response at different dose levels:
- Histologic (intra-alveolar oedema) evaluation after 12 h of instillation: Very slight, moderate and severe distribution of intra-alveolar oedema was observed at 1, 4 and 5 mg/kg bw respectively. Interstitial oedema was observed.
3. Results for investigation of dose level which produces early alveolar oedema and alveolar surface protein, after 20 h of instillation:
- Lung wet weight: Slight increase was observed, except for 0.25 mg/kg bw (not significant)
- Alveolar suface protein: Dose dependent increase, ranging from 5-50 mg, was observed
- Others: Upto 1.5 mg/kg bw, no visible signs of lung haemorrhage and red blood cells were observed in the lavage
4. Results for investigation of effects of perfusion procedure after 20 h: No significant changes in lung weight or in the alveolar surface protein were observed for perfused and non-perfused rat lungs
5. Results for investigation of time course of oedema and any signs of fibrogenesis: See Table 2
6. Results of histopatholgical examination of lungs at 6, 28, 72 h, 7 d and 14 d post-instillation: Interstitial fibrosis was observed in all the lungs after 14 d post-instillation

Any other information on results incl. tables

Table 1:Results forinvestigation of lung response at low dose (0.3 mg/kg bw) after 2 or 4 wks:

Time after Instillation (weeks)	Treatment	n	Lung wet wt (g) as mean± SD	Alveolar surface protein (mg/rat) as mean± SD
2	Distilled water	4	1.27 ± 0.08	3.5 ± 1.4
	Zinc chloride	4	1.27 ± 0.04	4.6 ± 3.9
4	Distilled water	4	1.47 ± 0.15	4.3 ± 1.4
	Zinc chloride	4	1.72 ± 0.32	34.4 ± 50.5

n = No. of animals

* = Significantly different from controls by Student's t-test for unpaired samples

Table 2: Results for effect of2.5 mg/kg bw zinc chloride on lungoedema and alveolar surface protein at different time intervals and lavaged lung dry weights, and hydroxyproline levels (indicator of fibrogenesis) after 7 and 14 d post-exposure

Treatment	Time after instillation	n	Lung wet weight (g) asmean± SD	Alveolar surface protein (mg) as  mean± SD	Dry lung weight (g) as  mean± SD	Hydroxyproline (mg/lung) asmean± SD
Control	6 h	6	1.29 ± 0.38	5.41 ± 2.43	-	-
Zinc chloride	6 h	6	1.97 ± 0.74	51.17 ± 40.09*	-	-
Zinc chloride	18 h	6	1.95 ± 0.34	30.09 ± 13.28	-	-
Control	28 h	4	1.10 ± 0.17	5.59 ± 2.46	-	-
Zinc chloride	28 h	6	2.80 ± 0.71*	71.97 ± 14.88*	-	-
Zinc chloride	2 d	5	2.21± 0.66	37.76 ± 25.85	-	-
Control	3 d	4	1.09 ± 0.04	3.25 ± 0.28	-	-
Zinc chloride	3 d	6	2.43 ± 0.64*	36.91± 1.76*	-	-
Control	7 d	4	1.13 ± 0.08	5.18 ± 2.08	0.188 ± 0.010	2.12 ± 0.26
Zinc chloride	7 d	6	2.05 ± 0.47*	9.99 ± 4.02	0.222 ± 0.023*	3.59 ± 0.88*
Control	14 d	5	1.17 ± 0.12	4.16 ± 0.69	0.166 ± 0.069	1.80 ± 0.25
Zinc chloride	14 d	6	1.57 ± 0.42	13.77 ± 7.83*	0.202 ± 0.069	3.46 ± 1.56

n = No. of animals

* = Significantly different from controls by Student's t-test for unpaired samples

Applicant's summary and conclusion

Conclusions:

Significant lung damage, in terms of oedematous reaction, was induced in rats on single intra-tracheal instillation of zinc chloride

Executive summary:

A number of single intra-tracheal instillation studies were performed in Wistar rats to investigate the delayed respiratory adverse-effects, and the potential of zinc chloride to damage the lungs.

 

In the study for investigation of pulmonary effects at low dose (0.3 mg/kg bw), a delayed elevated alveolar surface protein, with accompanying increased lung wet weight were observed after 4 wks. These effects correlated histologically with the quantitative biochemical assay for alveolar surface protein in a lavage fluid fraction.

Histopathological studies confirmed the induction intra-alveolar oedema at 12 h post-instillation of different dose levels (0.25-5 mg/kg bw) and interstitial fibrosis in the lungs at 2.5 mg/kg bw, after 14 d. The oedematous reaction in individual animals, after 20 h at 0.25-1.5 mg/kg bw, were variable, but the overall effect was dose dependent.  

 

The onset of oedema, was rapid with maximal effects being noted within 3 days when high doses (2.5 mg/kg bw) of test material were instilled. However, in majority of the animals the severe oedematous reaction regressed between 3 and 7 d. In some animals, adjudged by histochemistry and elevated lung hydroxyproline, evidence of a fibrogenic response was observed 7 d post-exposure.

 

Hence, significant lung damage, in terms of oedematous reaction was induced in rats, on single intra-tracheal instillation of zinc chloride.