Lambda 25 uv visible spectrophotometer

The antioxidant activity in the leaf extracts of Amaranthus hybridus was determined using the DPPH (diphenyl-picrylhydrazyl) and ABTS (2,2’-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid)) radical degradation technique.
DPPH assay: 10 μL of leaf extract is treated with 1 mL (250 μM) DPPH solution and incubated in dark at room temperature for 30 min. A change in color is estimated by recording the absorbance at 517 nm in a Perkin Elmer Lambda 25 UV/Visible spectrophotometer [39 (link)].
ABTS+ assay: Equal amount of ABTS (7.4 mM) and potassium persulfate (2.6 mM) were mixed and incubated at room temperature for 12 h in the dark. ABTS solution and the leaf extracts were mixed in a 19:1 ratio and incubated in the dark for 2 h. The absorbance against methanol was measured at 734 nm using a Perkin Elmer Lambda 25 UV/Visible spectrophotometer [40 (link),41 (link)].
The percent inhibition of DPPH and ABTS+ relative to the control was used to determine antioxidant activity using the following equation:
where Abs. blank—absorbance of the control 10 μL methanol for DPPH and 150 μL methanol for ABTS instead of Amaranthus hybridus leaf extract.
Abs. sample is the absorbance of the leaf extracts.
Ascorbic acid was used as a reference standard, and the results obtained are expressed as μg ascorbic acid equivalent g⁻¹ (μg AAE g⁻¹) dry weight.

The size distribution of sol particles was measured by a dynamic light scatter (Brookhaven, Holtsville, NY, USA, 90 plus). Infrared spectra were recorded on a PerkinElmer RXI FT-IR spectrometer (Waltham, MA, USA). Absorption spectra were measured with a Perkin-Elmer Lambda 25 UV-visible spectrophotometer. The microstructure of the sample was investigated by scanning electron microscopy (SEM, Hitachi, Tokyo, Japan, S-4800). Material thermal stability was measured by the thermogravimetry analysis (TGA, SDT-Q600, TA Instruments, New Castle, DE, USA). The refractive index of the optical films was measured by using the N & K analyzer (1280, n & k Technology, San Jose, CA, USA). The reflection and transmission spectroscopy were first measured and the data were calculated by using the Forouhi-Bloomer model to test the goodness of the fit. This process generates the values of refractive index (n) and extinction coefficient (k) which provide a basis of data to produce a new reflection curve through calculation. The curve fitting was repeated until this new reflection curve closes to the measured curve with the goodness of fit greater than 0.99. The obtained refractive index and extinction coefficient are reliable after the calculation and curve fitting. In the fitting process, the film thickness can simultaneously be obtained.

Quercetin was considered as the standard sample to compare the total flavonoid contents present in the plant extracts; 5% NaNO₃ was added to the leaf extract and mixed by vortexing. The mixture was allowed to stand for 6 min, following which, 10% AlCl₃ was added to the and incubated at room temperature for 5 min. Next, 1 M NaOH was added and the volume was made up to 5 mL with distilled water. The mixture was further incubated at 40 °C for 20 min, after which the absorbance was recorded at 430 nm using a Perkin Elmer Lambda 25 UV/Visible spectrophotometer [43 (link)]. The total flavonoid content (TFC) was calculated using the standard quercetin calibration curve, y = 0.605x + 0.0132, R2 = 0.9956 and was expressed in mg QE/g dry extract weight.