IMT-IAG-004E

Instruction Manual ofAgriChemTM* Pesticide Residue Detection Kit
TM*: Under registration in Japan

Harmful reagents are included. Please read this manual before use

Intended Use
AgriChemTM is designed for rapid test of residual pesticides (organophosphates and carbamates) in foodand environment without any measuring instrument. This kit is qualitative testing method and covers
most of organophosphates and carbamates and monitoring of pesticide drift.

Measurement Principle
Cholinesterase(ChE) is an enzyme that catalyzes the hydrolysis of the neurotransmitter acetylcholine
into choline and acetic acid, is an essential enzyme to allow a cholinergic neuron to return to its restingstate after activation , and is inhibited by organophosphates and carbamates.
In this kit, immobilized ChE to the bottom of microtube by hydrophilic resin is colored blue withsubstrate. When assay sample contains any organophosphates and carbamates, the top layer remains
white because of the ChE inhibition.

Agri
Chem

Add eachsolution

AgriChem

Positive
Before Test

Negative
After Test

Solid phasecholinesterase-1-

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Features1. Rapid: Compared with instrumental analysis and ELISA method, AgriChemTM is rapid testing.
2. Ease of use: AgriChemTM does not need any specific skills nor instruments and is testable even on site.
3. Low cost：Not necessary to use any detection equipment.
4. High sensitivity: More sensitive than conventional enzyme kits by column concentration.
5. Safety: AgriChemTM is designed for minimal exposure to reagents by performing reactions in tubes.

Components (contents)
Appearance (Below figure is the one for five tests. A part of bottles sizes change for other tests.

List
NameType

1. AgriChem
2. Solution-1（Blue cap）
3. Tube-2（Pink label）
4. Tube-3（Yellow label）
5. Solution-3（Yellow cap）
6. Solution-4（Red cap）
7. Color Index Card
8. Purified water（White cap）
9.Column tip
10.Syringe (10mL)
11.Filter pack12.Tube rack

Solution
PowderSolution
Solution
SolutionIAG-2020test
22
120
20
1
1
1
120
120
1

IAG-6060test66
36060
331260160
1For control reaction, AgriChem and Tube 3 are contained 1 more in 5-tests kits, 2 more in 20-tests kit,
and 6 more in 60-tests kit than number of tests.

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Assay Protocol
Materials required but not provided in kit box
1. Kitchen knife and chopping board
2. Plastic bags for samples
3. Scales in grams for samples（1g～500g）
4. 100mL beaker (or equivalent)
5. 200mL beaker (or equivalent)
6. Timer（in second）
7. City water (or purified water)
8. Waste tank

Preparation of the Reagents （Prepare below reagents before test）
Solution 2（Activator Solution）：Add 1ml (up to long line of picture (below))of Purified water into Tube 2. Shake and solve the white powder well (Solution 2).

２※Prepared Solution-2 is not possible to preserve nor re-use.

Adjust only the inspection numbers of thissolution and use once and daily. Do not re-use.

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Testing Procedures & Instructions
Testing of negative control needs to be run always at same time of each sample testing to make surethat the kit performs properly. There may be a case that the degree of color development changes byroom temperature for enzyme reaction. AgriChem for control test is not operated up to ⑰ after ①

Preparation and Extraction of Residual Pesticides in Crops ：Extraction of Adherent
Pesticides on the Surface of Crops.
① Prepare the tests number + 1 (for negative control) of AgriChem, open the caps, and apply 2 dropsof Solution 1 into AgriChem.

After applying, close the caps.

1AgriChem② Weigh the crop sample and put into a plastic bag. About 100g～300g of crop is proper to handle forelution (to be varied according to the type of crop). When the size and weight become problems, cutit in a suitable size with the cutting board and the kitchen knife.

③ Add an equal weight of water ※1（1g=1mL） into the bag by beaker （of 100mL）※1 If available, purified (distilled or deionized) water is recommended.

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④ Seal and shake the bag vigorously for 30 seconds for preparation of sample extraction.

⑤ Pour 100ml of the extraction water in the bag into beaker (of 100mL).

⑥ For removing solid contaminants in extraction water, set the filtration pack onto beaker (of 200mL)
and add whole extraction water to the filtration pack.

Collect more than 10ml of filtrated

extraction water and discard the filtration pack and the remaining extraction water on filter.

Adsorption and Activation of Pesticides ： Concentration and Activation of Pesticides
in Extract on Column.
⑦ Put the column tip firmly onto the top of 10mL syringe (Column Syringe).
Syringe

Column tip

担体

Syringe(10mL)

ColumnColumn tip

Column Syringe-5-

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⑧ Aspirate slowly※2 10mL of the filtrated extraction water from the beaker （of 200mL） into theColumn Syringe.

⑨ With the Column Syringe, pipette (dispense and aspirate) slowly※2 the extraction water into andfrom 100mL beaker. Repeat three times. At this step, pesticides in the extraction water are trappedon the resin on the top of column tip. Discard the extraction water in Column Syringe.

×3
終了後

Waste after
廃棄theend※2 Strong aspiration may cause detachment of the resin from the top of tip. Aspirate slowly.
Aspirate less than 1ml per 2 sec. and idle at every 1ml aspiration⑩ Set the tube of Solution 2 on tube lack, with the Column Syringe, pipette whole volume of theprepared Solution 2 from and into the Tube-2 once. (Organophosphorus pesticides adsorbed on thecolumn are converted to activated (oxon) form.)

２⑪ The remained activator solution in the Column Syringe needs to be completely drained by moving
the piston of

the syringe up and down

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⑫ Pour about 20 ml of fresh (city) water into beaker (of 100mL). Pipette 5mL of this water forwashing inside of the Column Syringe. If Solution 2 remained in Column Syringe, enzymatic
reaction is affected. Repeat 3 times with changing the fresh (city) water every time.

×3Exchange
Every time

⑬ The remained water in the Column Syringe needs to be completely drained by moving the piston ofthe syringe up and down.

Elution of Pesticides and Inhibition of Cholinesterase ： Transfer Pesticides fromColumn Syringe to AgriChemTM.
⑭ Pour 3 drops of Solution 3 into Tube 3 and set Tube 3 on tube lack.

3⑮ Pipette the whole volume of Solution 3 from and into the Tube 3 with the Column Syringe. It must
be cautioned that the liquid doesn't go up to the syringe※3.

The solutiondose not goover this line
at aspiration

３※3 For small amount aspiration, aspirate slowly while turning the piston of the syringe. Dispense
normally.
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⑯ Repeat step ⑮ three times. At this step, pesticides on resin are eluted into Solution 3.

３⑰ Set the tube of AgriChem on tube lack, aspirate the whole volume of Solution 3 from the Tube 3,and dispense the whole solution into AgriChem (for testing). Apply 3 drops of Solution 3 from
Bottle 3 in to AgriChem (for negative control). After closing the cap of AgriChem, shake lightlyand wait for 15 minutes for inhibiting enzymatic reaction.

３AgriChem

Color Development ： Confirmation of the Presence of Pesticides by the Enzyme
Reaction.
⑱ Apply one drop of the Solution 4 into AgriChem (for testing and negative control). After closing thecap of AgriChem, shake lightly and wait for 15 minutes. Handle Solution 4 carefully, because
Solution 4 is easily spilled from the bottle.

4AgriChem⑲ Compare the color of top layer of AgriChem with the Color Index CardAgriChem-8-Copyright (C) Institute of Microchemical Technology Co., Ltd.

Read Result
Compare with the top layer of resin (area of dotted circle-line in the leftpicture) and Color Index Card. If necessary, overlap AgriChem onto theColor Index Card.

Negative : The pesticide is not detected.
Positive, Marginal: The pesticide or enzyme inhibitor remains in thesample. If necessary, re-examine and confirm with instrumental test
method.

Examples of Detection Limit of Pesticides by AgriChemTM
with Standard Solutions
Pesticides
Detection Limit（ppb）
EPN60AZINPHOS-ETHYL
2AZINPHOS-METHYL
10ISOXATHION
0.6
ETHION20ETRIMFOS10QUINALPHOS
0.4
COUMAPHOS6CHLORPYRIFOS
0.4
CHLORPYRIFOS-METHYL
2CHLORFENVINPHOS40SULPROFOS
60
DIAZINON
0.8

Pesticides
TERBUFOSPARATHIONPYRIDAFENTHIONPIRIMIPHOS-METHYL
FENAMIPHOSFENITROTHIONPROPAPHOS
BROMOPHOS-ETHYLPHOSALONE
PHORATE
MONOCROTOPHOSPIRIMICARB

Detection Limit（ppb）
60
1220680
6208100
10010The expression “1 ppb” means that given property exists at relative proportion of one part per billionparts examined, as would occur if water-borne pollutant was present at concentration of one-nano gramper gram of sample solution (0.000001%).
Confirm the latest maximum residue limits (MRLs) of agricultural compounds in foods in each country.

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Storage and Stability
Store between 4℃ and 8℃. Protect from light. Do not freeze. The expiration date is indicated on box.

.

Transportation
Keep cool between 4℃ and 8℃. Protect from light. Do not freeze.

Cautions①For Safety・Do not drink and do not touch the reagents. This kit contains the harmful reagents. For use this kit,
treat carefully for safety by wearing lab coat, protection grove, and protection glass. If you feelabnormality in your body, see medical doctor immediately.
・After testing, AgriChemTM is possible for containing high concentration of pesticides. Do not open thecap of AgriChemTM after testing.

②For General
・This kit is designed for self-inspection related to food hygiene. Operate and judge the result at yourresponsibility.
･Waste the surplus reagents and inspection samples according to the regional regulations, underconsidering health and environment well. If AgriChemTM judged the positivity, there is a possibility
that pesticides exist in the AgriChemTM, and treat the AgriChemTM as polluted waste.
・Store this kit at 2-8℃ under the dark condition.
・Keep and refer to this kit manual carefully at testing and when necessary.
・In farm products, influence of crops origin materials and impurities might be received, and the false
positive can be caused.

Contact below address when such a result is doubted.

Contact Information
Further information on this product is available directly from Institute of Microchemical Technology.
Contact via website http://www.i-mt.co.jp/.
Any change of the specifications of this product may be applied without any advance notice.

Manufactured by:

Institute of Microchemical Technology
713A West KSP, 3-2-1 Sakado, Takatsu, Kawasaki, Kanagawa 213-0012, Japan
TEL：+81-44-811-6521/ FAX：+81-44-814-5545
http://www.i-mt.co.jp/- 10 -

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