Germicide Residual Test Result

Conducted by Japan Food Hygiene Association(JFHA), Tokyo, Japan, which is the Corporation Aggregate established and registered under authorization of Ministry of Health, Labor and Welfare based on the Japanese Laws of Japan Food Hygiene(Law No.233 from Dec 24, 1947) and The Pharmaceutical Affairs(Law No.145, from Aug 10, 1960)

E-Plan has commissioned JFHA to test the rate of residual of germicide using mini-tomatoes with TPN(chlorothalonil) applied on them on October 28th 2008.

TPN applied to tomatoes:
ST "DANIKORU"(ダニコール) 1000 manufactured by Sumitomo Gardening Chemical(住友化学園芸社)

Testing Method:
Gas Chromatography

Object tested "1"
Mini-tomato soaked in the city water with TPN 1,000x diluted for one minute and then dried in the normal temperature for 24 hours

Object tested "2"
Mini-tomato treated as "1" and then washed by normal city water for three minute

Object tested "3"
Mini-tomato treated as "1" was soaked in undiluted "E-Wash", pH12.5 alkaline water supplied by E-Plan and then washed by normal city water for one minute

Report No. T081-03945 dated Nov 11 2008 for Object "1"
Rate of residual of TPN: 4.3ppm

Report No. T081-03948 dated Nov 11, 2008 for Object "2"
Rate of residual of TPN: 3.5ppm

Report No. T081-03951 dated Nov 11 2008
Rate of residual of TPN: 2.0ppm



Human Skin Test Result

Test and Report made by Institute of General Health Development Co., Ltd.(SOUKEN), Tokyo, Japan requested by E-Plan Co. Ltd., Funabashi, Japan

1) Product Tested:
① High Alkali Water pH12.5 (RO water 99.9% Potassium Carbonate 0.1%) produced by E-Plan Co., Ltd (Equivalent to BANO)
② Saline (as control – comparative product)
③ White Vaseline (as control – comparative product)

2) Subjects (Examinees):
Total 22 with breakdown of Female 17 (age ranging from 22 to 56), Male 5 (age ranging from 37 to 55) who were recruited with full informed consent and the test was conducted under the ethical observation under the Helsinki Declaration"(Revised in Seoul 2008)

3) Test Method and How the Test was done:
Test was made in accordance with the method "HITO-PATCH TEST" – Human Patch Test - stipulated in "the guide book 2008 for manufacture and sale of cosmetic and relative non-medical products" to determine the level of stimulation to and the physical changes of the human skin.

After the physical examination of the subjects (examinees), the position of the skin on the back of each subject (examinee) was determined and picture by digital camera was taken.

0.03ml of ① ② ③ was applied to Patch Tester "Tori-i" manufactured by Torii Pharmaceutical Co., Ltd. and attached to designated position of each subject(examinee) and they went back home. Bathing and shower was prohibited while the patches were on. = June 21, 2010



After 24 hours, all subjects (examinees) were requested to come back to the Clinic and patches were taken off. At 60 minutes after patches were taken off, examination by doctor was made. Then the positions where the patches were on were marked and pictures were taken by digital camera. = June 22, 2010

All subjects (examinees) were requested to come back to the Clinic again at 24 hours after the patches were taken off and doctor’s examination and picture taking were done same as above. = June 23, 2010 (Camera used: Digital Camera Nikon D50 manufactured by Nikon K.K.)

Measurement and evaluation on the effect to the skins after the test were made also in accordance with stipulated process in the above guide book and evaluation was made by Takashi Koikeda, licensed doctor at Shiba Palace Clinic, Tokyo, Japan.

Evaluation was made under the following criteria:

Basis of Judgment Judgment Rating
No Reaction Negative(-) 0
Slight Stigma Slightly Positive (±) 0.5
Obvious Stigma Positive(+) 1.0
Stigma+Oedema+Papule Highly Positive(++) 2.0
Stigma+Oedema+Papule+Small Blister Highly Positive(+++) 3.0
Large Blister Highly Positive(++++) 4.0

Skin Stimulation Index = Total Rating of those subjects(examinees) who claims significant reaction to the skin / number of subjects(examinees) X 100

Below 5.0 Safe
5.0 – 15.0 Within Allowance
15.0 – 30.0 Improvement Necessary
Above 30.0 Hazardous

After careful examination and evaluation based on the above procedures and criteria, All products ① ② ③ were rated "0" including Skin Stimulation Index and all products are confirmed to be completely safe.

Signed,

Koikeda Takashi
Doctor in Charge of Examination and Evaluation



Effect of Antibacterial Test Result - Test 1

To: E-Plan Co., Ltd.
August 10, 2009
Institute registered under the law of public hygiene of Japan
Public Health Foundation of Miyagi Prefecture
Chief Director Sakuo Matsuzaki

Based on your request for testing the performance of ionized alkaline water in terms of effect of antibacterial, the result is reported as follows:

Product supplied for testing: Ionized alkaline water pH12.5
Bacteria roots supplied: Salmonella arizonae ATCC13314
Number of n: n=3

Testing Method:
(1) Produce of bacteria liquid:
Supplied root of bacteria was inoculated to normal vegetable gelatin and cultured for 2 times for 24 hours under the temperature of 35°C. Also supplied root of bacteria was cultured at NB medium for 24 hours under the temperature of 35°C. Both cultured salmonella arizonae bacteria was diluted by sterilized saline solution to make bacteria contained liquid to be viscosity of 10³ - 10⁴ /ml as object of testing.

(2) How testing was made:
100μl of liquid bacteria produced as above (1) was added to 10ml of pH12.5 ionized alkaline water and left for 30 seconds, 5 minutes and 15 minutes under the temperature of 20°C. Each specimen was filtered by membrane of 0.45μm and washed by sterilized purified water until the specimen does not show any alkalinity. Membrane after filtering the specimen was attached to the base of normal vegetable gelatin and cultured for 2 days under the temperature of 35°C. Then the number of bacteria after culturing was counted and number of bacteria per 1ml was calculated. The same process was conducted using Sodium hypochlorite as control.




Effect of Antibacterial Test Result - Test 2

To: E-Plan Co., Ltd.
August 10, 2009
Institute registered under the law of public hygiene of Japan
Public Health Foundation of Miyagi Prefecture
Chief Director Sakuo Matsuzaki

Based on your request for testing the performance of ionized alkaline water in terms of effect of antibacterial, the result is reported as follows:

Product supplied for testing: Ionized alkaline water pH12.5
Bacteria roots supplied:
① Escherichia coli ATCC8739
② Vibrio parahaemolyticus ATCC17802
③ Salmonella anatum ATCC9270
④ Escherichia coli O157 ATCC35150
Number of n: n=3

Testing Method:
(1) Produce of bacteria liquid:
Supplied roots of bacteria of the above ①③④ were inoculated to normal vegetable gelatin and cultured for two times for 24 hours under the temperature of 35°C. Also supplied roots of bacteria was cultured at NB medium for 24 hours under the temperature of 35°C. Both cultured bacteria ①③④ as such were diluted by sterilized saline solution by 10 times to make bacteria contained liquid to be of viscosity of 10⁶ - 10⁷ /ml as object of testing. As to ②, bacteria root was treated in the same manner as for ①③④, but cultured at normal vegetable gelatin with 3% volume content of sodium chloride, NB medium with 3% volume content of sodium chloride and diluted by sterilized saline solution of 3% volume content and to make bacteria contained liquid to be 10⁶ - 10⁷ /ml as object of testing.
(2) How testing was made:
100μl of liquid bacteria ①③④ produced as above were added to 10ml of pH12.5 ionized alkaline water and left for 30 seconds, 5 minutes and 15 minutes under the temperature of 20°C. Each specimen was filtered by membrane of 0.45μm and washed by sterilized purified water until the specimen does not show any alkalinity. Membrane after filtering the specimen was attached to the base of normal vegetable gelatin and cultured for two days under the temperature of 35°C. Then the number of bacteria after culturing was counted and number of bacteria per 1ml was calculated. As to ②, Membrane after filtering was attached to the base of normal vegetable gelatin with sodium chloride added by 3% and washing was done with normal city water with sodium chloride added by 3%. Number of bacteria was counted in the same manner for ①③④. As control, sterilized city water was used instead of pH12.5 ionized alkaline water.




Effect of Antibacteria Test Result - MY CO2 SDN BHD

Conducted by MY CO2 Sdn Bhd, an independent laboratory which provides analytical testing services internationally.

Product : Bano Super Alkaline Ionized Water
Bacteria : ① Salmonella Enterica (ATCC 14028)
  ② E-Coli (ATCC 25922)
  ③ Staphylococcus Aureus (ATCC 33862)
  ④ Vibrio Parahaemolyticus (ATCC 17802)
Contact Time : 30 Seconds, 5 Minutes and 15 Minutes
Effect of Antibacteria
Bacteria Initial count of test
microorganism per ml
Count of surviving test
microorganism per ml
Reduction Percentage kill
of test
microorganism
Cfu / ml Log 10 Cfu / ml Log 10
Salmonella Enterica
(ATCC 14028)
4.1 x 108 8.6 < 10 1.0 7.6 99.9999
E-Coli
(ATCC 25922)
3.3 x 108 8.5 < 10 1.0 7.5 99.9999
Staphylococcus Aureus
(ATCC 33862)
2.7 x 108 8.4 < 10 1.0 7.4 99.9999
Vibrio Parahaemolyticus
(ATCC 17802)
1.7 x 108 8.2 < 10 1.0 7.2 99.9999





Capability to Remove Radioactive Substances

News dated June 14, 2011

Regarding the capability of high alkaline ionized water to remove radiation contaminants or radioactive substances

This testing was made by Japan Functional Food Analysis and Research Center in Fukuoka, Japan to test how much high alkaline water produced by the electrolyte ionizing machine of E-Plan ( identical to the machine operating at Metita Sdn Bhd in Malaysia) can remove radioactive substances adherent to vegetables.

Based on the fundamental knowledge that high alkaline water can remove various agricultural chemicals attached to vegetables without any help of chemical detergent as high alkalinity works as surfactant and with the belief that the same function could be found in removal of radioactive particles / substances on the vegetable, the testing was made as follows:

Spinaches were taken from the farmer's house at Iidate Village in Fukushima Prefecture located very close to the exploded Fukushima Nuclear Power Plant No.1 and examined radiation dosage of the spinaches without washing and after washing with high alkaline ionized water.

The details of testing method and the results are as follows:
1. The date and time when the spinaches as specimen were taken: March 30, 2011, 1:00pm

2. The location the specimen spinaches were taken: Farm land near by Aza-Sawamegi, Kusano, IIdate Village, Fukushima Prefecture

3. Specimen taken: Spinaches 1 kg.

4. Washing method: 500 grams of spinaches were soaked in high alkaline water of pH12.5 for 5 seconds and washed by 500cc of RO water.

5. Testing Institute: Japan Functional Food Analysis and Research Center, 3-23, Tenya-cho, Hakata-ku, Fukuoka Prefecture following the testing process chapter 2-2 from "Food Radiation Contaminants Monitoring Manual under Emergency" issued by Safety Monitoring Section of Food Health Preservation Department, Medicine Bureau of Ministry of Health, Labor and Welfare of Japan using Nuclide Analysis Method by Gamma Ray Spectrometer and Germanium Semi-Conductor Detector.

6. The date of testing: April 7th, 2011

7. Test Result: (Unit: Bq/kg = Bq is Becquerel – International radiation measurement unit)

Items examined Not washed Washed Removed % Removal
Radioactive Iodine131 24,900 14,200 10,700 43%
Radioactive Cesium 187,400 31,900 155,500 83%
Cesium 134 88,600 16,100 72,500 82%
Cesium 137 98,800 15,800 83,000 84%

Observation:
Iodine 131 was removed only by 43% while Cesium was removed by more than 80%. This removal ratio will go higher if vegetables are soaked in alkaline water longer and also the radioactive dosage of spinaches as taken from the farm land was extremely high.

In reality, the temporary food safety criteria set by Japanese government for Iodine 131 is 2000Bq/kg and if vegetables with radiation level of 3000Bq/kg is washed by high alkaline water, the remaining will be 1710Bq/kg(3000X0.57=1710) which will be under the safety criteria.

Safety criteria for Cesium set by Japanese government is 500Bq/kg and if 1000Bq/kg vegetables are washed as above and calculated based on the above test result, the remaining is 170Bq/kg(1000X0.17=170)

It is safely understood that alkaline water with pH as high as 12.5 can remove almost all of radioactive contaminants if washed longer and rinsed more thoroughly than done in the testing.

Note: This test result is applicable only to high alkaline water produced by Metita Sdn Bhd and not to any other alkaline water in general.




Food Standard Test Result - MY CO2 SDN BHD

Conducted by MY CO2 Sdn Bhd, an independent laboratory which provides analytical testing services internationally.
Sample tested : Bano - Super Alkaline Ionized Water

Analysis Report
Test Description Analysis Result Standard Method Maximum Permitted

Physical
pH 12.0 APHA 4500-H+ B 6.5 - 8.5
Turbidity * 0.2 NTU APHA 2130 B NMT 5NTU
Color < 5 Pt Co APHA 2120 B NMT 15 Pt Co

Microbiological
Coliform ND (<1 cfu / 100ml) APHA 9222 B (2005) Must be absent
E-coli ND (<1 cfu / 100ml) APHA 9222 B (2005) Must be absent

Chemicals
Chloride (as Cl) ND < 2 mg/L APHA 4500-Cl-D (2005) NMT 250 mg/L
Fluoride (as F) ND < 0.02 mg/L APHA 4500-F-D NMT 1.5 mg/L
Magnesium 0.04 mg/L APHA 3111 B (2005) NMT 150 mg/L
Sodium (as Na) 17.0 mg/L APHA 3111 B (2005) NMT 200 mg/L
Sulphate (as SO4) ND < 0.3 mg/L APHA 4500-SO42-D (2005) NMT 400 mg/L
Anionic Detergent (MBAS) * 0.02 mg/L APHA 5540 C (2005) NMT 1.0 mg/L
Biocides (Total) * ND < 0.1 mg/L Biocides test kit NMT 0.1 mg/L
Carbon Chloroform Extract * ND < 0.01 mg/L Combustion infrared NMT 0.5 mg/L
Chromium (as Cr) ND < 0.01 mg/L APHA 3111 B (2005) NMT 0.05 mg/L
Chloroform * ND < 0.02 mg/L Headspace/ GC-FID NMT 0.03 mg/L
Cyanide (as CN) ND < 0.02 mg/L HACH Spectrophotometer Method 8027 NMT 0.1 mg/L
Mineral Oil ND < 0.2 mg/L APHA 5520 B (2005) NMT 0.3 mg/L
Phenol ND < 0.001 mg/L Manual UDK126 D & APHA 5530-D (2005) NMT 0.002 mg/L
Selenium (as Se) ND < 0.001 mg/L APHA 3500-Se D (2005) NMT 0.01 mg/L
Silver (as Ag) * ND < 0.01 mg/L APHA 3111 B (2005) NMT 0.05 mg/L
Zinc (as Zn) ND < 0.01 mg/L APHA 3111 B (2005) NMT 5.0 mg/L
Lead (as Pb) ND < 0.01 mg/L APHA 3111 B (2005) NMT 0.05 mg/L
Mercury (as Hg) ND < 0.001 mg/L APHA 3112 B (2005) NMT 0.001 mg/L
Arsenic (as As) ND < 0.001 mg/L APHA 3500-As C (2005) NMT 0.05 mg/L
Cadmium (as Cd) ND < 0.001 mg/L APHA 3111 B (2005) NMT 0.005 mg/L
Copper (as Cu) ND < 0.01 mg/L APHA 3111 B (2005) NMT 1.0 mg/L
Aluminium (as Al) 0.10 mg/L APHA 3500-Al D (2005) NMT 0.2 mg/L
Manganese (as Mn) ND < 0.01 mg/L APHA 3111 B (2005) NMT 0.1 mg/L
Iron (as Fe) ND < 0.02 mg/L APHA 3111 B (2005) NMT 0.3 mg/L
Ammonia (as N) * 0.17 mg/L APHA 4500 NH3 F NMT 0.5 mg/L
Hardness (as CaCO3) ND < 4 mg/L APHA 2340 C NMT 500 mg/L
Nitrate (as N) 0.30 mg/L APHA 4500-NO2 B (2005) NMT 10 mg/L
Residual Chlorine 0.04 mg/L HACH Spectrophotometer Method 8021 NMT 0.1 mg/L

Radioactivity
Gross α < 0.01 Bq/L Direct measurement using geiger meter 0.1 Bq/L
Gross β < 0.01 Bq/L Direct measurement using geiger meter 1.0 Bq/L

***ND = Not Detected





Virus Inactivation Test Result

Japan Food Research Laboratories

Virus Inactivation Test
1 Client

ePLAN

2 Specimen material

e WASH pH12.5

3 Trial Objective

Inactivating examination of Feline Calicivirus

4 Test Overview
Created an Action Liquid by adding and mixing a virus suspension of feline calicivirus to a specimen. Allowing it to set at room temperature, we measured the viral infection value of the action liquid after 30 seconds and again after 2 minutes, then 5 minutes. We then compare these results against the preliminary examination as a method of measurement of the vital infection value.
The feline calicivirus is used widely as a substitute for the norovirus from which cell cultures cannot be derived.

5 Test Results
1) Preliminary Examination
It confirmed that there was a viral infection value by diluting action liquid by a factor of 10 in a cell maintenance nutrient medium without being affected by the specimen.

2) The measurement of the viral infection value
The result is shown in List-1.
List-1     Viral infection value result of a measurement of the action liquid
Examination
Virus
Objective log TCID 50/mL *1
Start 30sec. later 2min. later 5min. later
*2
Feline calicivirus
Specimen 6.0 < 1.5 < 1.5 < 1.5
Contrast 6.0 - - 5.7

TCID 50 : median tissue culture infections dose, 50% tissue culture infectious [infective] dose
Start time : Measured TCID 50 of the contrast right after the action started
Contrast : Purified water
Action temperature : Ambient Room Temperature
- : Not Conducted
< 1.5 : Not Detected
*1 Numerical value of TCID 50 per 1mL of active liquid
*2 Substitute virus replacing the Norovirus

6 Testing Method
1) Test virus
Feline Calicivirus F-9 ATCC VR-782

2) Used Cell
CRFK cell [Dai Nippon Pharma]

3) Culture Medium
① Cell Growth Medium
Added 10% cow embryo serum to the eagle MEM nutrient medium "NISSUI".
② Cell Maintenance Medium
Added 2% cow embryo serum to the eagle MEM nutrient medium "NISSUI".

4) Preparation of Virus Suspension
① Cell Culture
Using a cell proliferation nutrient medium, a monolayer culture of a cell was obtained in a flask for tissue culture(s).
② Inoculation of the Virus
Removed cell proliferation nutrient medium from the flask after monolayer culture, then inoculated the examination virus. Added a cell maintenance nutrient medium, then cultivated it for 1~5 days in a carbon dioxide incubator (5% of CO2 density) of 37 degrees Celsius ± 1°C.
③ Preparation of the virus suspension
After incubations, observed the formation of the cell using a phase contrast inverted microscope. Then, confirmed that alteration of cellular morphology (cytopathic effect) was produced.
And centrifugalize (3000 r/min. ten minutes) of culture fluid, and virus suspension liquid was provided from the supernatant solution.

5) Test operation
Produced action liquid by adding and mixing 0.1mL of virus suspension into 1mL of liquid. Stored it at room temperature and measured a viral infection value of the action liquid diluting by a factor of 10 with cell maintenance nutient medium after 30 seconds, then 2 and 5 minutes.
Then, examined it in the same way using purified water as contrast, and measured at the beginning then 5 minutes later.

6) The measurement of the Viral Infection Value
After having monolayer culture of a cell in a microplate (96 well) for tissue culture with cell proliferation nutrient medium, removed cell proliferation nutrient medium and then added a cell maintenance nutrient medium of 0.1mL. Next, provided 10 times serial dilution of action liquid using a cell maintenance nutrient mediium after diluting the contrast 10 times.
Inoculated 0.1mL of diluted solution into by 4 holes and cultivated it for 4~7 days in a carbon dioxide incubator (5% of CO2 density) with 37 degrees Celsius ± 1°C.
After incubations, observed the formation of the cell using a phase contrast inverted microscope in order to check whether the alteration of cellular morphology (cytopathic effect) was occurring or not.
Then, calculated quantity of 50% tissue culture infection (TCID 50) by the Reed-Muench method and converted it into a viral infection value per 1mL action liquid.


Period.

foundation
Japan Food Research Laboratories