METHODOLOGY

Study Area

Borno State is located between latitude 10N and 13N and longitude 12 and 15. It is in the
north-easterly region of Nigeria, with an area of 70, 898 km2. Borno State is divided into 27 local
government areas (LGAs) with a total population of 4, 171, 104 [9]. The three seasons observed are: the
cool dry (harmattan) season (OctoberMarch), hot dry season (April-June) and rainy season (July-
September). Temperatures are generally high all the year round, with hot season temperatures ranging
between 39C and 40C under the shade. Average annual rainfall is between 500mm and 800mm. The
variability of rainfall is over 100 per cent. The relative humidity is generally low throughout the state,
ranging from as low as 13% in the driest months of February and March to the highest values of 78% in
the rainy season months of July and August [10].

Sampling

Samples of Beans (Phaseolus vulgaris) were collected in both field and storage facilities in six
Local council areas (LGA): Bama Biu, Damboa, Gubio, Maiduguri Metropolitan, and Monguno, in Borno
State. Sampling was conducted in two phases. Firstly, during the harvest periods (November - December
2008) from eighteen different farms (three each at the six LGAs) and secondly, six months post harvest
period at storage and preservation facilities. The post harvest samples consists of samples not limited to
the last farm harvests. Ten kilograms composites of each sample were collected in sterile poly bags on
location basis.

Determination

The determination of pesticide residue followed the multiresidue pesticide analysis technique
consisting of the QuEChERS (Quick, Easy, Cheap, Effective, Rugged and Safe) sample preparation method
by AOAC Method 2007.01 [11] with slight modifications for number of analytes spiking was adopted in
the work.

Extraction

A thoroughly homogenized 15 g sample of beans was weighed into an extraction tube. Then, 15
mL of 1% acetic acid-MeCN extraction solvent was poured into the tube on top of the sample. The
surrogate and the pesticide solutions were spiked into this MeCN layer for the method validation (MVD)
and method detection limit (MDL) samples. The tube was capped and vortexed for 30 seconds. The cap
was removed and the powder reagents were poured slowly into the MeCN layer. The cap was tightened
securely on the 50-mL extraction tube and was vortexed for 30 seconds until all of the powder reagents
were mixed with the liquid layers. The tube was placed on a mechanical shaker for 5 min and then
centrifuged for 5 min at 3, 000 rpm. Following this, 11 mL of the top MeCN layer were removed and
transferred to a 15-mL clean-up tube. This tube was capped and vortexed for 30 seconds and
centrifuged for 5 min at 3,000 rpm. A 5-mL aliquot of the top layer was transferred into a clean test tube
for solvent exchange.
Solvent Exchange

The 5-mL aliquot of the cleaned-up extract was blown down to dryness with a gentle stream of
nitrogen at 40 C in approximately 1 hour. Care was taken to remove the tube immediately when dried.
A 900-L aliquot of hexane/acetone (9:1) was added and 100 L of the internal standard (d10-parathion)
was spiked into the organic solution. The tube was capped and vortexed for 15 seconds. Next, 1 mL of
extract was transferred to a 2-mL clean-up tube, capped tightly and vortexed for 30 seconds. After
centrifuging for 5 min at 3,000 rpm, 200 L of the clear extract were transferred to an auto-sampler vial
with a small glass insert for analysis on the GC-ion trap MS. The individual calibration levels were spiked
into each extract for the calibration curve in matrix before the final clean-up step.

GC/MS Injection, Separation and Detection

The SHIMADZU GC/MS (GC-17A), equipped with fluorescence detector was used for the
chromatographic separation and was achieved by using a 35% diphenyl/65% dimethyl polysiloxane
column. The oven was programmed as follows: initial temperature 40 C, 1.5 min, 25 C/min to 150 C,
0.0 min, 5 C/min to 200 C, 7.5 min, 25 C/min to 290 C with a final hold time of 12 min and a constant
column flow rate of 1 mL/min. The detection of the pesticides was performed using the GCion trap MS
with optional MSn mode. This scanning mode offered enhanced selectivity over either full scan or
selected ion monitoring (SIM). The GC-ion trap MS was operated in the MSn mode and performed
tandem MS functions by injecting ions into the ion trap and destabilizing matrix ions, isolating only the
pesticide ions. The retention time, peak area and peak heights of the sample were compared with those
of the standards for quantization.

Data Analysis

Results obtained were analysed on Microsoft Excel 2007 spreadsheet.