Phosphine

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1990

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

GLP compliance:

yes

Type of assay:

bacterial gene mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Phosphine

EC Number:

232-260-8

EC Name:

Phosphine

Cas Number:

7803-51-2

Molecular formula:

H3P

IUPAC Name:

phosphane

Test material form:

other: gas

Method

Target gene:

Histidine loci

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium TA 1538

Species / strain 2

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102

Metabolic activation:

with and without

Test concentrations with justification for top dose:

See tables in results section.
Range covered = 4.52 - 4340 ppm
One original study followed by 4 re-tests

Vehicle / solvent:

not applicable, mixed with air to achieve ppm

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

air

Positive controls:

yes

Positive control substance:

9-aminoacridine

2-nitrofluorene

sodium azide

mitomycin C

other: 2-anthramine, 2-aminofluorene, ethylene oxide, 1,3-butadiene

Details on test system and experimental conditions:

Cultures were obtained from Dr. Bruce N Ames, Berkely, CA and stored frozen.
Fresh cultures for mutagenesis testing were prepared by quickly thawing a vial of frozen working stock cultures of each tester strain and transferring the culture to 25 ml of Oxoid Nutrient Broth #2. After growth for approximately 10 hours at 37°C in an orbital shaking incubator, samples of each culture were diluted 1:4 in distilled water and optical densities 0.40 to 0.60 (approximately 1-2 x 109 cells/ml; representative of cells in late exponential or early stationary phase) were utilized for this study.
Treatments were performed by combining 2 ml top agar (supplemented with O.5mM histidine/O.5mM biotin), 0.1 ml tester strain and 0.1 ml control article (if appropriate) in sterile glass tubes preheated to 45°C. The tubes were vortexed and the mixture was poured onto minimal glucose plates, evenly distributed, and allowed to solidify. Cultures treated in the presence of S9 also contained 0. 5 ml of the S9 mixture. Within an hour the untreated and "standard" positive control plates were inverted and incubated in the dark at 37°C for 48 hours. Cultures treated by exposure to gaseous or volatile test or control articles were prepared as for the untreated cultures. However, these cultures were placed, inverted without lids, in 250-nun glass desiccators (approximate volume 5L). All strains with and/or without S9 receiving a particular treatment were combined in a single desiccator. Desiccators receiving PH3 or l,3-butadiene were sealed prior to introduction of approximately 6-8 volumes the gaseous test or control articles.
To evaluate contamination plates wihtout Salmonella cultures were also scored for growth.

Evaluation criteria:

Scoring
Following the 48-hour incubation and 24 hour de-gas periods, the background lawn and spontaneous revertants were scored for normal, inhibited or no growth. Inhibited growth was characterized by the absence of a confluent bacterial lawn and/or the presence of pindot colonies. Revertant colonies were enumerated on an Artek electronic colony counter interfaced with an IBM PC/AT computer for data acquisition. Solvent and positive controls were scored first, and test article treated cultures were scored only if the average negative control values approximated historical ranges (x ± 2SD).
Evaluation Criteria
A positive result is defined as a statistically significant, dose-dependent increase in the number of histidine-independent revertants with at least one dose level inducing a revertant frequency that is two-fold the spontaneous solvent control value. Statistical analyses were performed using the program developed by Snee and Irr (1981), with significance established at the 95% confidence limit. If the test article does not induce a statistically significant, dose-dependent increase in revertant frequency but does induce a revertant frequency at one dose level that is two-fold the spontaneous control value, the result is considered equivocal. A negative result is defined as the absence of a statistically significant or dose-dependent increase in the number of histidine-independent revertants.

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at some dose levels

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 1538

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at some dose levels

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 3

Species / strain:

S. typhimurium TA 102

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

The increases observed in strains TA1535, TA1537, TAl.538 or TA98 in the first three assays were never independently confirmed and are considered to be statistical aberrations or artifactual in nature. Therefore, the results for PH3 were negative in the Ames/Salmonella Plate Incorporation Assay under theconditions, and according to the criteria, of the test protocol.

Any other information on results incl. tables

See method section for original test and first re-test (note not all numbers easy to read on original report)

Mean of triplicate values shown. Shading = cytotoxicity

 

treatment	dose (ppm)	S9	TA1535	TA1537	TA1538	TA98	TA100	TA102
Retest 2
untreated		-	12	6	5	15	105	208
air		-	8	8	6	18	95	295
Phosphine	99	-	11	6	5	21	94	294
	152	-	11	5	4	21	91	275
	183	-	15	5	5	17	95	295
	111	-	10	6	3	18	103	275
	172	-	10	5	7	16	96	228
	262	-	11	4	4	23	101	205
untreated		+	11	9	5	23	101	272
air		+	10	6	14	26	93	321
Phosphine	99	+	6	7	17*	27	108	290
	152	+	6	9	13*	29	124	265
	183	+	5	8	16*	25	103	276
	111	+	5	9	19*	35	112	279
	172	+	7	6	16*	30	113	262
	262	+	4	6	20*	31	97	206
treatment	dose (ppm)	S9	TA1535	TA1537	TA1538	TA98	TA100	TA102
Re-test 3
untreated		-	8	9	10	22	68	305
air		-	9	8	11	15	72	256
Phosphine	37	-	8	5	5	23	78	223
	99	-	11	5	3	19	74	206
	75	-	12	5	6	25	89	239
	138	-	10	5	5	14	93	79
	143	-	16*	5	5	23	80	128
	203	-	14	5	4	20	88	92
untreated		+	8	11	16	30	88	391
air		+	8	9	10	27	62	265
Phosphine	37	+	11	7	19	33	84	254
	99	+	9	8	22	31	94	229
	75	+	9	8	17	26	86	270
	138	+	10	8	16	23	92	75
	143	+	8	6	20	34	103	135
	203	+	13	7	15	26	86	74
treatment	dose (ppm)	S9	TA1535	TA1537	TA1538	TA98	TA100	TA102
Re-test 4
untreated		-	14	8	10	34	91	365
air		-	15	6	9	32	81	307
Phosphine	41	-	13	7	10	35	98	251
	139	-	12	5	6	34	96	69
	277	-	10	4	6	15	49	1
	518	-	3	2	1	13	8	1
	670	-	1	2	1	7	3	0
	962	-	2	0	1	4	0	0
untreated		+	13	12	16	41	101	329
air		+	17	12	16	33	92	404
Phosphine	41	+	11	6	16	39	96	286
	139	+	13	8	16	42	111	90
	277	+	9	5	12	12	25	15
	518	+	8	1	3	3	14	5
	670	+	3	5	8	8	8	7
	962	+	4	1	1	1	9	1

 

 

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative with metabolic activation
negative without metabolic activation

Executive summary:

Phosphene did not induce gene mutations in the plate-incorporation Ames test in any of the strains tests, with and without metabolic activation.