Genesis 10s uv vis

Uronic acid was determined using the method of Blumenkrantz and Gustav (1973). Briefly, 1 mL ion-exchange water was added to each sample, and 1 mL iced concentrated sulfuric acid (0.025 M borax) was mixed into 200 µL of each sample. After heating in 100 °C water for 10 min and cooling in ice, 40 µL carbazole solution (125 mg carbazole, 100 mL ethanol) was mixed into each sample. The samples were heated in 100 °C water for 15 min, then cooled in ice; absorbance was measured at 530 nm (GENESIS 10S UV-VIS; Thermo Scientific). The cell-wall material was treated for 4 h at 4 °C with 0.1 M NaOH to saponify the methyl and acetyl esters. The suspensions were adjusted to pH 5.0 with 10% (vol/vol) glacial acetic acid and then treated for 16 h at 30 °C with a homogeneous preparation of endo-polygalacturonase (EPG) from Aspergillus niger [2.5 units, Megazyme, Wicklow, Ireland; 1-unit releases 1 μmol of reducing sugar min⁻¹ from a 1% (weight/volume) solution of polygalacturonate at pH 5.0 and 25 °C. The suspensions were centrifuged, and the insoluble residues were washed with water.

The synthesis of SeNPs was confirmed by observing absorption peak maxima between 200 and 500 nm in a spectrometer (Genesis 10S UV–Vis, Thermo Scientific, Carlsbad, USA). The contribution of possible functional groups of A. glaucum leaf extracts to the synthesis of SeNPs was evaluated by FTIR analysis (Nicolet iS 5, Thermo Scientific, Waltham, USA) in the 4000-400 cm⁻¹ region. Morphological characterization of SeNPs was performed by SEM (MIRA 3 LMU, TESCAN, Brno, Czech Republic) at 20 kV and TEM (JSM-1010, JEOL, Tokyo, Japan) at 80 kV. To determine the size distribution of SeNPs dispersed in the aqueous medium, the DLS technique was performed using a Zetasizer (Nano-ZS90, Malvern Instrument, United Kingdom). The elemental composition of the SeNPs was determined by energy dispersive X-ray spectroscopy (EDS) using a microprobe for surface microanalysis (Quantax EDS, Bruker, Billerica, Germany).

The cultures sampled (50 mL) were centrifuged for 10 min at 3500 rpm in VWR Mini Star microcentrifuge (VWR, Radnor, PA, USA). The supernatant was collected to quantify glucose, phosphate, and ammonium concentrations.
When necessary, the supernatant was diluted in a saline solution (10% sodium chloride, 90% distilled water). Freestyle precision Neo kit (Abbott, Witney, Oxon, UK) was used to determine glucose concentration in g/L.
Ammonia and phosphate Sera Tests (Sera, Heinsberg, Germany) were used to determine ammonium and phosphate concentrations, respectively. The supernatant was diluted with distilled water when necessary. The absorbance was measured at the wavelength of 697 nm for ammonium and 716 nm for phosphate. The absorbances were measured using Genesis 10S UV–Vis (Thermo Scientific, Waltham, MA, USA).

The AE was determined in triplicate, using an indirect method after separation by centrifugation of the particles from the aqueous medium containing the non-associated insulin. The amount of insulin associated with nanoparticles was calculated by measuring the difference between the total amount of insulin added to the formulation, and the insulin released in the supernatant. Quantification of insulin was performed according to the Bradford method [28 (link)], and using bovine serum albumin (BSA) as standard protein. Absorbance was read using a spectrophotometer (Genesis 10S UV-vis Thermo Scientific), at 595 nm. The AE was determined applying the following equation: $\mathrm{AE}(\%)=\frac{Totalamountofinsulin-insulininsupernatant}{Totalamountofinsulin}\times 100$

Uronic acid was determined using the method of Blumenkrantz and Gustav (1973) (link). Briefly, 1 mL ion exchange water was added to each sample, and 1 mL iced concentrated sulfuric acid (0.025 M borax) was mixed into 200 µL of each sample. After heating in 100 °C water for 10 min and cooling in ice, 40 µL carbazole solution (125 mg carbazole, 100 mL ethanol) was mixed into each sample. The samples were heated in 100 °C water for 15 min, then cooled in ice; absorbance was measured at 530 nm (GENESIS 10S UV-VIS; Thermo Scientific).

Transesterification activity of recombinant lipase Lk2 and Lk3 were measured using Pohnlein method with slight modification [21 (link)]. Transesterification was performed between methyl esters and para-nitrophenol in organic solvents at various temperature. The conversion of para-nitrophenol into para-nitrophenyl esters was monitored through decreasing absorbance at 400 nm. The reaction procedures as followed: 10 µl concentrated recombinant lipase was mixed with 990 µl organic solvents (contained 4 mM pNP and 24 mM methyl esters). In a 2 ml reaction tube, the reaction was carried out for 10 min at 150 rpm. Enzymes precipitation were done by centrifuged the tube at 11,900 g for 3 min, then 50 µl of the upper layer was removed and mixed with 1 ml of Tris Cl buffer pH7 (0.1 percent triton). UV-Vis spectrophotometry was used to measure the remaining pNP (Genesis 10S UV-Vis, Thermo Scientific). The amount of enzyme that released one mole of p-nitrophenyl ester in 1 min was defined as one enzymatic unit. All of the tests were carried out in triplicate, including the blanks (without the addition of enzymes).

The determination of the absorption spectrum and the stability of the SeNPs were obtained using a UV-vis spectrophotometer (Genesis 10S UV-vis, Thermo Scientific, Carlsbad, CA, USA) in the wavelength range from 200 to 400 nm with a path length of 1 cm. The size and shape of the SeNPs were determined by SEM (MIRA 3 LMU, TESCAN, Brno, Czech Republic) at 20 kV and TEM (JSM-1010, JEOL, Tokyo, Japan) at 80 kV. A descriptive statistical analysis was performed on the size distribution of SeNPs from three independent assays. A mean of 53.30 nm, median of 48.72 nm, mode of 48.72 nm, standard error of 3.49, as well as Kurtosis coefficient (0.1011) and skewness coefficient (0.8387) close to zero were obtained. From these data, the normal distribution plot (Gaussian distribution) of the SeNPs size was plotted.Surface microanalysis microprobe EDX spectroscopy (Quantax EDS, Bruker, Billerica, MA, USA) was used to detect the elemental composition of the SeNPs. The functional groups present in the extracts and the resulting SeNPs were determined by FTIR spectrophotometry (iS10 FTIR, Thermo Scientific, Waltham, MA, USA).