Method development and validation for the quantitative determination of total flavonoids through the complexation of iron (III) and its application in real sample.

BACKGROUND: The determination of flavonoids in real sample using UV-Vis spectrophotometer commonly uses quercetin and catechin with Al+3 complexing agent as reference materials for the calibration of the instrument. However, getting these standard materials is challenging due to its expense and unavailability in the chemical reserve of the country. Moreover, the Al+3 - quercetin complexation standard method demands high amount of quercetin in spite of its high cost. Hence, developing alternative method that can solve this problem is crucial for the determination of flavonoids in the real sample. RESULTS: An iron-based complexation method for the determination of flavonoids in the real sample was developed that reduces the amount of quercetin by 200 times (1 mg/mL to 0.005 mg/mL) during the calibration of UV-Vis spectroscopy as an alternative method. The reaction parameters (incubation time, pH, and concentration of quercetin) were optimized using software Design Expert 11 and confirmed by the practical experiments. The kinetics of reaction between iron and quercetin was found to be pseudo first order with rate constant of kobs at 340 and 510 nm. The analysis window for the flavonoid complex was achieved with the kinetic discrimination of the interferences at its optimized time of complexation 20 min and absorbance maxima of 510 nm. The developed method was validated by evaluating its precision, accuracy, recovery test (84-117%), detection limit and quantification limit following the standard protocols. The calibration of the instrument has been developed for the new method and the linear regression coefficient (R2) of 0.998 was obtained. SIGNIFICANCE: Applying the developed standard material (Fe3+ - quercetin complex) gives freedom for the analytical chemists to find the standard materials that is accessible and cheaper than the existing one (Al3+-quercetin complex). The developed method can also be easily applied for determination of flavonoid in the real samples without potential interferences coming from sample matrix.

Anchote (Coccinia abyssinica) starch extraction, characterization and bioethanol generation from its pulp/waste.

A polysaccharide molecule called starch exists in nature and is cheap, renewable, biodegradable, and readily accessible. The main objective of this project is to extract and characterize anchote (Coccinia abyssinica) starch, as well as to generate bio-ethanol from its pulp/waste. The anchote sample used in this investigation came from Ethiopia's western Oromia region. Anchote (Coccinia abyssinica) starch was extracted, and the pulp from the tuber was peeled and the supernatant isolated for bio-ethanol production. The extracted starch from anchote and bio-ethanol from pulps were characterized by physicochemical, functional properties and FT-IR analysis. The result of physicochemical and functional properties of anchote starch was found to be pH (4.44), WHC (112%), Solubility (5.03%), swelling power (5.781%), Gelatinization temperature (53.33 °C), WAC (2 g/g), bulk density (0.605 g/cm3) and OAC (3 g/g). According to this FTIR research, anchote starch has a chemical structure that is similar to corn, cassava, and potato starch. As the results showing a promising alcoholic content (25% v/v) and the existence of bioethanol being validated by a combustion test, the pulps/wastes recovered from anchote are also good sources of bioethanol. Finally, the FTIR spectroscopic analysis revealed that ethyl alcohol was produced from anchote pulps/waste after acid hydrolysis, fermentation, and distillation.

Evaluation of Proximate, Phytochemical, and Heavy Metal Content in Black Cumin and Fenugreek Cultivated in Gamo Zone, Ethiopia.

Spices have been recognized to have medicinal properties. Among several spices and medicinal plants, black cumin and fenugreek are very common. Geographical, climatic, and cultivating soil type can change the chemical composition of these spices. The objective of this work is to determine proximate, phytochemical, and heavy metal content in black cumin and fenugreek cultivated in the Gamo zone, Ethiopia. The proximate composition (moisture, ash, fiber, fat, protein carbohydrate, and gross energy) was determined; their content (%) in black cumin is 6.98 ± 0.12, 5.02 ± 2.9, 6.90 ± 0.21, 32.1 ± 0.68, 20.9 ± 0.09, 34.1 ± 0.73, and 498 ± 4.4, respectively, and 6.30 ± 0.35, 4.27 ± 0.17, 9.36 ± 0.25, 12.8 ± 0.41, 30.8 ± 0.09, 46.1 ± 0.52, and 422 ± 1.6, respectively, in fenugreek. The total means of phenolic flavonoids and alkaloids in black cumin are 193 ± 5.3 mg GAE/100 g, 87.6 ± 4.3 mg QE/100 g, and 69.4 ± 4.7 mg AE/100 g, respectively, and 382 ± 11 mg GAE/100 g, 123 ± 3.6 mg QE/100 g, and 37.6 ± 2.2 mg AE/100 g, respectively, in fenugreek. In black cumin and fenugreek collected from Kamba, Daramalo, and Dita woredas, nine heavy metals were determined by using FAAS. The total mean concentrations in mg/kg for detected heavy metals (Fe, Zn, Cu, Mn, and Co) in black cumin are 78.5 ± 5.9, 27.3 ± 1.5, 3.06 ± 0.26, 8.47 ± 0.22, and 10.1 ± 0.37, respectively, and Cr, Ni, Pb, and Cd were not detected in black cumin. Similarly, the concentrations of Fe, Zn, Cu, Mn, Co, and Ni in fenugreek are 168 ± 5.2, 14.8 ± 0.20, 4.76 ± 0.1, 13.7 ± 0.40, 3.66 ± 0.21, and 2.12 ± 0.03, respectively, and Cr, Pb, and Cd were not detected in fenugreek. All the results of determined parameters were compared to previous studies, and the values were in agreement with slight variations. Therefore, black cumin and fenugreek cultivated in the Gamo zone were free from heavy metal toxicity.