Ultraviolet 1800 spectrophotometer

All reagents used in synthesis and biological evaluation were commercial (Merck, Fischer and Acros Organics) and were used as supplied. Initial thiosemicarbazones (1) are readily available either commercially and via simple one-step literature procedures (see ref. 17–25 (link)). Chromatographic silica gel 60 (220–240 mesh) was obtained from Merck. Thin layer chromatography was performed on foil-backed plates coated with Merck Silica gel 60 F₂₅₄. Plates were developed using ultraviolet light and basic aqueous potassium permanganate. Uncorrected melting points were measured on a DMP-300 A&E Lab apparatus. Ultraviolet (UV) absorption spectra were recorded on Shimadzu Ultraviolet-1800 spectrophotometer in the range 200–800 nm. Infrared spectra were recorded on a Bruker OPUS FT-IR spectrometer by attenuated total reflection (diamond-ATR) on solid films. Proton and ¹³C NMR spectra were recorded on Bruker DPX-400 and 500 MHz spectrometers at ambient temperatures; where necessary HSQC and HMBC techniques were used to confirm structural connectivities. Mass spectra were recorded on Bruker Micro TOF-ESI positive targeted mode. Elemental (CHN) analyses were conducted on a CE-440 Elemental Analyzer.

All reagents
and solvents used
in syntheses of compounds 3a–o were
of analytical grade and used as received without further purification.
The reagents and solvents were purchased from commercial sources (Sigma-Aldrich
Merck, Fischer, and Acros Organics). Precoated thin-layer chromatographic
aluminum sheets (Kiesel gel 60, F₂₅₄, E. Merck, Germany)
were used to monitor the progress and purity of synthesized compounds.
Melting points (M.p.) were recorded in open capillaries using a DMP-300
A&E Lab, U.K., melting point apparatus and are uncorrected. Ultraviolet
(UV) absorption spectra were recorded on a Shimadzu Ultraviolet-1800
spectrophotometer in DMSO. IR spectra were recorded on a Bruker OPUS
using attenuated total reflectance (ATR) to identify the functional
groups. Proton (¹H), carbon (¹³C), and fluorine
(¹⁹F) NMR experiments were performed on Bruker DPX-400
and 500 MHz spectrometers. Mass spectra were recorded on the Bruker
Micro time of flight-electrospray ionization (TOF-ESI) positive targeted
mode. Fluorescence of glycated products was measured on a Shimadzu
RF-6000 spectrofluorometer.

The total phenolic content of the extracts was determined by the Folin–Ciocalteau method with some modifications.[13 (link)] 5 g/50 ml of sample was filtered with whatman number 1 paper. To 0.5 ml of the sample, 2.5 ml of 0.2 N Folin–Ciocalteau reagent was added and kept at room temperature for 5 min. After that, 2 ml of sodium carbonate (75 g/l) was added to reaction mixture and the total volume was made up to 25 ml using distilled water. The solution was then kept for incubation at room temperature for 2 h. Absorbance was measured at 760 nm using 1 cm cuvette in ultraviolet - 1800 spectrophotometer (Shimadzu, Kyoto, Japan). All procedures were performed with three replicates. Tannic acid (0.1–0.5 mg/ml) was used to produce a standard calibration curve. The correlation equation constructed with tannic acid was y = 1.633X (R² = 0.985). Total phenolic content was expressed as milligrams of tannic acid equivalents per gram of dry extract (mg TAE/g).