Diodon hystrix, commonly known as spot-fin porcupine fish is a salt-water fish belonging to the Diodontidae family. It is widely distributed in Sabah wet market due to its commercial value. This study exploits the effectiveness of Cholinesterase (ChE) obtained from the brain tissue of D. hystrix in detecting carbamates inhibitory activities. Carbamate pesticides known to inhibit ChE and toxic towards living organisms can contaminate the water bodies. By using diethylaminoethanol (DEAE) Sepharose ion exchange chromatography, a total of 40% recovery yield of ChE was obtained with a 165.77 purification fold. Furthermore, the ChE showed a high affinity towards acetylthiocholine iodide (ATC) with an optimum activity at pH 7.45 and temperature ranging from 20 to 40℃. Among five different types of carbamates, methomyl was found to have the highest percentage of inhibition analyzed using ChE inhibitory assay, followed by carbofuran, bendiocarb, carbaryl and propoxur with >85% inhibition rate. The results concluded that ChEs extracted from the brain tissue of D. hystrix are applicable to be used as a bioindicator in detecting the presence of carbamates.

The discharge of industrial effluents into nearby water bodies affects the inhabitants including living organisms. The presence of foreign materials such as heavy metals can be a threat to the ecosystem as they are enormously carcinogenic even though in minute concentration. Hence, an economical and time-efficient preliminary screening test is crucial to be developed for the detection of heavy metals, prior to employment of high technology instruments. In this study, cholinesterase (ChE) from Sabah porcupine fish, Diodon hystrix was purified to test for its potential as an alternative biosensor in detecting metal ions. Few enzymatic parameters including specificity of substrate, temperature and pH were applied to determine its optimal enzymatic activity. ChE enzyme was found to be more sensitive towards the presence of substrate, butyrylthiocholine iodide (BTC), in contrast to acetylthiocholine iodide (ATC) and propionylthiocholine iodide (PTC) with the effective coefficient at 7193, 3680.15 and 2965.26 Vmax/Km, respectively. Moreover, the extracted ChE enzyme showed the optimum activity at pH 9 of 0.1 M Tris-HCl and at 25°C to 30°C range of temperature. When subjected to heavy metals, ChE enzyme was significantly inhibited as the enzyme activity was reduced in the sequence of Hg > Ag > Cr > Cu > Cd > Pb ≥ Zn > As. As a conclusion, the partially purified ChE enzyme proved its sensitivity towards metal ion exposure and can be used as an alternative method in screening the level of contamination in the environment.

Cholinesterase-based biosensor well known as a sensitive method to detect the existence of harmful dissolved compounds in any type of water source, especially the river. This alternative biosensor can be used to determine the level of pollution of the water in a short period of time as well as to evaluate the low cost and simple service. The aim of this study was to exceed the effectiveness of acetylcholinesterase source extracted from the brain tissue of Asian swamp eel; Monopterus albus as a potential environmental biosensor. Purified acetylcholinesterase exposed to a different type of metal ions and mercury showed the highest percentage of inhibition at 62.9% followed by chromium at 59.22% while silver, arsenic, cadmium, cobalt, copper, nickel, zinc a¬¬nd lead at not more than 50% (approximately 37-50%). Metal ions such as mercury, zinc, chromium and copper showed exponential decay type inhibition curves with calculated half maximal inhibitory concentration; IC50 in the ascending sensitivity order 0.005, 0.595, 0.687 and 1.329 mgL-1, respectively. Field trial works exhibited that the acetylcholinesterase was applicable in sensing heavy metals pollution from the river which closed to the industrial and agricultural sites at near real-time and verified using ICP-OES. This study proves the potential use of acetylcholinesterase sourced from M. albus as a biomonitoring tool to assess the contamination level of the river.

Affinity chromatography for acetylcholinesterase; AChE namely Procainamide Sepharose CL-6B was synthesised through the coupling method between soluble procainamide hydrochloride immobilised by a cross-linked agarose size exclusion, sepharose CL-6B. 1, 4-butanediol diglycidyl ether plays a role in building up a productive and rigid connecting of a biospecific ligand (Procainamide hydrochloride) to an insoluble matrix (Sephacryl CL-6B). Local freshwater eel brain was extracted and centrifuge at high speed. The supernatant was collected, and seven different volumes were separately loaded on to the column for isocratic purification where 12 fractions were collected at the end of elution stage. This study shows 1000 μL of extractant; considered as the maximum volume to load onto the column. Lastly, a stepwise elution was performed with five different concentrations of NaCl, and each of 1 mL fraction was collected then assay for determination of enzyme activity and protein content. The data shows AChE was successfully purified with percentage recovery of 38 % after 21 purification fold. Kinetic study strengthens the data where the efficient coefficient ratio of ATCi was much higher compared to PTCi and BTCi at 3.03, 2.67 and 1.52 Vmax⋅Km −1, respectively, prove that the collected fraction predominantly contained with AChE, which is a targeted enzyme to be used as a sensitive biosensor to detect the presence of carbamate and organophosphate contamination in the environment.