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Gaches Chimie Report 3

Test report n° RE21-0054-2 Laboratoire MIDAC – 85 C, rue Nelson Mandela – 59120 LOOS/LILLE – FRANCE Tél : +33 (0)3 20 90 52 55 – contact@midac-lab.fr SAS au capital de 80 000 € - R.C.S. LILLE Métropole 329 303 242 Evaluation of the virucidal activity according to the NF EN 14476 + A2 : 2019 standard Product(1) : Global cold washing process, ozone generator OTEK with the addition of dedicated detergents (Lavage n°5: Lavage de type blanc souillé) Tested virus : Human coronavirus 229E (HCOV-229E) Test temperature : 10°C Batch(1) : / On request of(1) : GACHES CHIMIE SPECIALITES ACTIVITE ENTRETIEN TEXTILE 2 BIS CHEMIN DE LA SCIERIE FR 64800 OS MARSILLON Loos, the 25 February 2021 The test report includes : 9 pages
LABORATOIRE MIDAC RE21-0054-2 Page 2 sur 9 I. PRINCIPLE The virucidal activity was determined according to the protocol of the NF EN 14476+A2 standard: "Chemical disinfectants and antiseptics - Quantitative suspension test for the evaluation of virucidal activity in the medical area – Test method and requirements (phase 2, step 1)." - July 2019. II. IDENTIFICATION OF SAMPLE Product(1): Processus global de lavage à froid, Générateur à ozone avec adjonction de produits lessiviels dédiés (Lavage n°5: Lavage de type blanc souillé). Machine OTEK(1) Cool Star – Batch : 190422801 – Manufacture date : 29/08/2019 – Expiry date : / Hypochlorite de sodium 12.5% (47/55° chlorométrique) – Batch : 4200401 – Manufacture date : / – Expiry date : / Garosive deter – Batch: 200080001 – Manufacture date : 24/02/2020 – Expiry date : / Garo boost – Batch : 200101601 – Manufacture date : 05/03/2020 – Expiry date : / Peracid forte – Batch : 200015501 – Manufacture date : 15/01/2020 – Expiry date : / (Mix of products made by the MIDAC Laboratory with the addition of products in the vertical direction from top to bottom) Society: GACHES CHIMIE SPECIALITES Received at the laboratory: 17 June 2020 Storage conditions at the laboratory: Room temperature, in the darkness. Appearance of the product: Garosive deter is green liquid and others products are colorless liquid. Product diluent recommended by the manufacturer: Undiluted.
LABORATOIRE MIDAC RE21-0054-2 Page 3 sur 9 III. EXPERIMENTAL CONDITIONS Period of analysis : 18 January 2021 to 29 January 2021 Product test concentrations : see table(s) on next page(s). Virus identification: Human Coronavirus 229E (HCOV-229E), P1 Name and source : HeLa , CCL-2 Number of passages : 16 Cell culture medium : MEM 10%SVF, 1% AANE, 1%ATB, 1%L-Glu Product diluents used : Distilled water. Appearance product dilutions : Essay 1 : Colorless homogeneous liquid Essay 2 : Whitish homogeneous liquid Essay 3 : Whitish homogeneous liquid Stability of mixture: None precipitate observed during the test Test temperature : 10°C (± 1°C). Contact times : 7 minutes (+/- 10 secondes) Interfering substances : 3g/l bovine albumin with 3mL/L sheep erythrocytes (dirty conditions) Method used for product neutralization : Microfiltration method - Columns: Microspin S-400 HR Incubation temperature : 37°C (± 1°C) et 5 % CO2 IV. RESULTS The calculation of titer reductions is based on the method of Spearman and Kärber. 1. Method validation Assays were validated by: - a cytotoxicity control - a sensitivity control of untreated cells - a formaldehyde internal standard control - a product neutralization control
LABORATOIRE MIDAC RE21-0054-2 Page 4 sur 9 a) Product cytotoxicity Product cytotoxicity (log DICT50) Dilution method 4.50E+00 Filtration on Microspin TM S-400 HR columns 1.50E+00 Comment: after dilution according to the standard protocol, the cytotoxicity of the product is 4.50E+00 log DICT50. The filtration on Microspin TM S-400 HR columns leads to the reduction of the toxicity at 1.50E+00 log DICT50. b) Cells sensitivity Sensitivity (log DICT50) Treated cells A 5.38E+00 Untreated cells B 5.50E+00 Comment: The difference between the virus titer of treated cells (A) and untreated cells (B) must be less than 1 log. c) Formaldehyde internal standard control Product Test concentration Interfering substance Viral titer in the test (log DICT50) Reduction (log DICT50) Formaldehyde – 30 min 0.7% (v/v) PBS 3.13E+00 2.00E+00 Formaldehyde - 60 min 0.7% (v/v) PBS 2.75E+00 2.38E+00 Viral control formaldehyde n.a PBS 5.13E+00 Comment: The difference between the titer of the viral control, expressed as a logarithm, and the titer of the virus in the formaldehyde internal standard control is 2.00E+00 log DICT50 after 30 min and 2.38E+00 log DICT50 after 60min.
LABORATOIRE MIDAC RE21-0054-2 Page 5 sur 9 d) Neutralization control Product Test concentration Interfering substance Viral titer in the test (log DICT50) Reduction (log DICT50) Neutralization of the product Essay 3 3g/l bovine albumin with 3mL/L sheep erythrocytes (dirty conditions). 5.63E+00 0.00E+00 Neutralization control n.a 5.63E+00 Comment: Checking the efficiency of the neutralization of product activity, the difference of titer with the test suspension must be ≤ 0.5log. 2. Test results: Evaluation of virucidal activity Product Test concentration Interfering substance Viral titer in the test (log DICT50) Reduction (log DICT50) Global cold washing process, ozone generator OTEK with the addition of dedicated detergents Lavage n°5: Lavage de type blanc souillé: / Essay 1 3g/l bovine albumin with 3mL/L sheep erythrocytes (dirty conditions). 3.88E+00 1.75E+00 Essay 2 1.50E+00 4.13E+00 Essay 3 1.50E+00 4.13E+00 Viral control T0 5.63E+00 Viral control Tmax 5.63E+00 Comment: At least one concentration per test must show a reduction of 4log or more, and at least one concentration must show a log reduction of less than 4.
LABORATOIRE MIDAC RE21-0054-2 Page 6 sur 9 V. CONCLUSION According to NF EN 14476+A2: 2019 standard, The product Global cold washing process, ozone generator OTEK with the addition of dedicated detergents (Lavage n°5: Lavage de type blanc souillé) (1) batch(1) : / possesses a virucidal activity tested at Essay 2 after 7 minutes (+/- 10 seconds) at 10°C (± 1°C) with 3g/l bovine albumin with 3mL/L sheep erythrocytes (dirty conditions) against Human Coronavirus 229E, (HCOV-229E). VI. REMARKS The COFRAC accreditation attests laboratories are competent for the only tests covered by the program. Copy of this test report is authorized only in its entirety. This report concerns only the tested product. (1) Data provided by the customer. Laboratoire MIDAC assumes no responsibility for the information provided by the customer VII. REVISION Date Revision description Version n.a n.a n.a
LABORATOIRE MIDAC RE21-0054-2 Page 7 sur 9 Annex 1 : Raw Data Product(1) : Global cold washing process, ozone generator OTEK with the addition of dedicated detergents (Lavage n°5: Lavage de type blanc souillé) Batch(1) :/ Product Concentration Interfering substance Contact Time 10-1 10-2 10-3 10-4 10-5 10-6 10-7 10-8 Global cold washing process, ozone generator OTEK with the addition of dedicated detergents (Lavage n°5: Lavage de type blanc souillé) Cytotoxicity Essay 3 3g/l bovine albumin with 3mL/L sheep erythrocytes (dirty conditions) n.a TTTT TTTT TTTT TTTT TTTT TTTT TTTT TTTT 0000 0000 0000 0000 0000 0000 0000 0000 Global cold washing process, ozone generator OTEK with the addition of dedicated detergents (Lavage n°5: Lavage de type blanc souillé) Cytotoxicity after Microspin Essay 3 n.a TTTT TTTT 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 Sensitivity of treated cells Essay 3 60min 3333 3333 2222 2222 1111 1111 1110 1111 1101 1101 1010 0000 0000 0000 0000 0000 Sensitivity of untreated cells n.a PBS 60min 4444 4444 2222 2222 2122 2223 3122 2222 1001 1133 1010 0000 0000 0000 0000 0000 Formaldehyde 30min 0,7 % (v/v) 30min 4444 4444 4444 4444 1100 1034 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 Formaldehyde 60min 0,7 % (v/v) 60min 4444 4444 4444 4444 0001 0001 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 Formaldehyde control n.a 60min 3333 3333 3333 3333 1111 1233 0111 0133 1100 0033 0100 0011 0000 0000 0000 0000 Formaldehyde Cytotoxicity 0.7%(v/v) n.a 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 Viral control n.a 3g/l bovine albumin with 3mL/L sheep erythrocytes (dirty conditions) 0 2222 2222 1111 1111 1111 1123 0010 1112 1111 1111 1100 0011 0000 0000 0000 0000 Viral control n.a 7 minutes (+/- 10 seconds) 2222 2221 1111 1111 1111 1111 1111 1102 1111 1001 1001 1001 0000 0000 0000 0000 Global cold washing process, ozone generator OTEK with the addition of dedicated detergents (Lavage n°5: Lavage de type blanc souillé) Essay 1 7 minutes (+/- 10 seconds) 1111 1111 1011 1111 1111 1110 1001 1101 0000 0000 0000 0000 0000 0000 0000 0000 Essay 2 4444 4444 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 Essay 3 4444 4444 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 Inactivation Essay 3 30min 4444 4444 4444 4444 2222 2222 2111 1111 1101 1111 0100 0100 0000 0000 0000 0000 Inactivation control n.a 30min 2222 2222 1111 1111 1111 1111 1111 1111 1110 0111 0110 0010 0000 0000 0000 0000 1 to 4 Presence of virus. 1 = 25% CPE and 4 = 100% CPE 0 Absence of virus T Toxic n.a non-applicable
LABORATOIRE MIDAC RE21-0054-2 Page 8 sur 9 Annex 2: Graphic presentation of the test results a) Reference inactivation of the virus by 0.7% (v/v) formaldehyde b) Evaluation of virucidal activity of the product 5,631,501234560 min7 minlg DICT50Contact time Cytotoxicity of the tested productInactivation of the virus by the tested product5,133,132,750123456030min60minlg DICT50Contact timeInactivationof the virus by 0,7%(v/v) formaldehydeCytotoxicity of 0,7% (v/v) formaldehyde
LABORATOIRE MIDAC RE21-0054-2 Page 9 sur 9 Annex 3: Methodology verification A test is only valid if the following criteria are fulfilled: a/ Test virus suspension has at least a concentration which allows the determination of a 4 log10 reduction of the virus titer. b/ The difference between the titer of the viral control, expressed as a logarithm, and the titer of the virus in the inactivation reference test is between -0,5 and -2,5 after 30min and between -2 and -4,5 after 60min for poliovirus, between -3 and -5 after 30min and -3,5 and 5,5 after 60min for adenovirus, between 0.0 and -2 after 30min and -0.5 and 2,5 after 60min for parvovirus, between -0.75 and -3,5 after 5min and between -2,0 and ≥ -4 after 15min for vaccina virus, between -1 and -3 after 30min and between -2,0 and -4,0 after 60min for murine norovirus c/ Cytotoxicity of the product does not affect cell morphology and growth or susceptibility for the test virus in the dilutions of the test mixtures which are necessary to demonstrate a 4log reduction of the virus. d/ Comparative titration of the virus on the treated cell cultures with dilutions of the test mixture and in parallel with PBS results in a difference <1 lg of the viral titer. e/ When checking the efficiency of the neutralization of product activity, the difference of titer with the test suspension must be ≤ 0.5log. f/ At least one concentration per test must show a reduction of 4log or more, and at least one concentration must show a log reduction of less than 4.