RBPR technology uses the enzyme acetylcholinesterase (AchE) isolated from housefly brains to detect organophosphate and carbamate pesticide residues through an enzymatic reaction. In Nepal, a RBPR laboratory was established in 2014 to test agricultural products for pesticide residues. The AchE test involves mixing the enzyme with chopped produce samples - a slowed or stopped reaction indicates the presence of insecticides. Results are quantified within 15 minutes to determine if the sample is safe for consumption, requires quarantine, or should be considered not edible based on the level of inhibition observed. While effective, the AchE test can sometimes produce false negatives by not detecting all types of insecticides.

1. RAPID BIOASSAY OF
PESTICIDE RESIDUE (RBPR)
TECHNOLOGY
Prepared by:
Dev Bhattarai
Agriculture and Forestry University
Rampur Chitwan, Nepal

2. RBPR technology is developed by Taiwan Agriculture
Research Institute (TARI) in 1958 to check the residue of
organophosphate and carbamates groups of pesticides and
fungicides residue in certain level through enzymatic
reaction.
In Nepal, RBPR laboratory is established in Kalimati on 18
June 2014 under the supervision of Plant Protection
Directorate.
Chemical used in this technology is AchE (Acetyl
cholinesterase) isolated from the brain of House fly.

3. When AchE reaction solution is mixed with samples
from healthy produce, it slowly turns yellow. If
insecticides is present the enzymatic reaction slows
down or stops.
The rate at which color development is inhibited
indicates the quantity of chemicals present, as well as
their toxicity.
The AchE test is very rapid, taking only 15 minutes
from sampling to completion.
The B.t. test uses Bacillus thuringiensis to detect
fungicide residues.

4. B. thuringiensis is a bacterium which is entomopathogenic i.e.
it causes disease only in insects and is highly sensitive to some
fungicides used on vegetables.
Vegetable samples are finely chopped and placed in EDTA
solution for 5 minutes. The extract is incubated with a
suspension of B. thuringiensis for 90 minutes (the period of four
generations of bacteria). Then TTC solution is added and left to
act for another 30 minutes. Finally, the reaction is halted with
Triton-HCl mixture, which acts as a red color signal for B.
thuringiensis activity.

5. If fungicide residues are present, there is no metabolic activity
by the bacteria and no red color. If by the end of the test period
the mixture has not turned red but is still its original pale
yellow, it probably contains toxic chemical residues.
Garlic and ginger can kill bacteria naturally, and can’t be tested
by B.t.. Some compounds which occur naturally in oranges and
tea also interfere with the TTC reaction. Strawberry fruits
contain a red pigment, which makes it difficult to distinguish
any color change from bacteria. It is possible to test these two
groups, but the procedure must be modified.

6. The result withdrawn from the inhibition percentage given
by AchE test:
Below 35%: edible (Green)
35-45%: Need quarantine for few days (Yellow)
More than 45%: not edible (Red)
The inhibition % is calculated through
Spectrophotometer. The increasing absorbance for
sample at 412 nm is recorded, compared to the
insecticide free blank (normal) AchE reaction and the

7. % inhibition= Absorbance(normal)–Ab. change(sample) *100
Absorbance change (normal)
The AchE test doesn’t give false positive results i.e. it doesn’t
indicate insecticides are present when they are not. However, it
does sometimes give a false negative. It can’t detect certain
insecticides, for example, synthetic pyrethroids, and gives a
negative result even if they are present.
References: Rapid Bioassay-FFTC Publication Database
An Overview of Pesticide Management in Nepal-The journal of Agriculture and Environment vol:18-
Adhikari Parashu Ram