Cary 8454 uv vis spectrophotometer

Manufactured by Agilent Technologies

Sourced in United States

The Cary 8454 UV-Vis spectrophotometer is a laboratory instrument designed to measure the absorption or transmission of light in the ultraviolet and visible regions of the electromagnetic spectrum. It is capable of analyzing a wide range of samples, including liquids, solids, and gases.

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15 protocols using cary 8454 uv vis spectrophotometer

Kinetics of Array Formation Analyzed

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Arrays formation kinetics was determined by turbidity due to light
scattering, monitored by absorption at 330 nm wavelength, using an Agilent
Technologies (Santa Clara, CA) Cary 8454 UV-Vis spectrophotometer. Absorption
spectrum at wavelengths range of 190 nm to 1100 nm was acquired every 5 seconds
for 25 minutes immediately following an initial blanking. Absorption curves at
330nm were constructed of measurements of blank samples (buffer: 25 mM Tris-HCl,
150 mM NaCL, 5% glycerol, and 500 mM imidazole) as control, Bcomponents at 5 μM, and A+B mixtures (5, 10 or
15 μM). Curves were acquired for three experimental replicates for each
experimental condition (two for blank control). Curves were processed as
follows: the respective initial value (first time point) was first subtracted
from each curve to account for initial background; then, a non-linear offset was
applied by subtracting the averaged curve of the blank measurements from each
and all the other curves. Extended Data Figure
5a shows the average absorption of each group of samples and standard
deviation (n=3 experimental replicates). All data was processed using python
Dataframe and Numpy packages.

Ben-Sasson A.J., Watson J.L., Sheffler W., Johnson M.C., Bittleston A., Somasundaram L., Decarreau J., Jiao F., Chen J., Mela I., Drabek A.A., Jarrett S.M., Blacklow S.C., Kaminski C.F., Hura G.L., De Yoreo J.J., Ruohola-Baker H., Kollman J.M., Derivery E, & Baker D. (2021). Design of Biologically Active Binary Protein 2D Materials. Nature, 589(7842), 468-473.

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Photoactivated Pigment Kinetics Analysis

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All of the experiments were carried
out at pH 7. The UV–vis absorption spectral measurements were
taken with a Cary 8454 UV–vis spectrophotometer (Agilent Technologies,
CA) equipped with a thermostated cuvette holder (Agilent 89090A).
All of the light reactions at various temperatures were performed
under an identical irradiation condition with a Schott 250 W cold
light source (with a long-pass (>520 nm) cutoff filter). The temperature
fluctuated by ±0.5 °C.
The concentration of hydroxylamine
used (0.5 M) in the reactions was considerably high relative to the
protein concentration (which was in the range of 1.5–2 μM);
thus, the reaction can be assumed to be of first order with respect
to the pigment. Therefore, the kinetic traces were followed at the
pigment absorption maxima and fitted to single-exponential or biexponential
decay components by the following equation or
where y is the pigment remaining, k is the rate of the reaction, and a and b are the coefficients related to the relative amount of
each component. To determine the activation energy (E_a) and the frequency factor (A) of the
processes, the obtained reaction rates in the dark and in light at
various temperatures (T) were fitted to the Arrhenius
equation.

Das I., Pushkarev A, & Sheves M. (2021). Light-Induced Conformational Alterations in Heliorhodopsin Triggered by the Retinal Excited State. The Journal of Physical Chemistry. B, 125(31), 8797-8804.

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DPPH Antioxidant Activity Assay

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The DPPH free radical technique was used to calculate the AOA. The extracts’ free radical scavenging abilities against the DPPH free radical were determined spectrophotometrically using the method reported by Mustafa et al. [19 (link)].
To 4.5 mL of 0.1 mM DPPH (ethanolic solution), 0.5 mL of extract solution was added. After mixing, the solution was stored in the dark for 30 min at room temperature before the disappearance of DPPH color was spectrophotometrically measured at 517 nm with an Agilent Cary 8454 UV-Vis spectrophotometer. The results were represented as mg Trolox equivalent (TE)/g dry weight (DW) using Trolox as the reference antioxidant.

Mustafa A.M., Abouelenein D., Acquaticci L., Alessandroni L., Abd-Allah R.H., Borsetta G., Sagratini G., Maggi F., Vittori S, & Caprioli G. (2021). Effect of Roasting, Boiling, and Frying Processing on 29 Polyphenolics and Antioxidant Activity in Seeds and Shells of Sweet Chestnut (Castanea sativa Mill.). Plants, 10(10), 2192.

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Quantifying Polyphenol Content in Food

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Total phenolic content was determined following the Folin–Ciocalteu
(FC) method based on the work by Kosar et al. (2005), and proanthocyanidin
content was determined according to DMAC, vanillin, and butanol/HCl
assays used by Montero et al. (2013), Gu et al. (2008), and Pérez-Jiménez
et al. (2009), respectively, employing a Cary 8454 UV–vis spectrophotometer
(Agilent Technologies, Palo Alto, CA, USA).^{29 (link),44 (link)−46 (link)} The results were expressed as milligrams of epicatechin
per 100 g of sample.

Domínguez-Rodríguez G., Ramón Vidal D., Martorell P., Plaza M, & Marina M.L. (2022). Composition of Nonextractable Polyphenols from Sweet Cherry Pomace Determined by DART-Orbitrap-HRMS and Their In Vitro and In Vivo Potential Antioxidant, Antiaging, and Neuroprotective Activities. Journal of Agricultural and Food Chemistry, 70(26), 7993-8009.

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Characterizing P22 Virus-Like Particles

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10 μL of P22 VLPs (10 mg/mL in water) was added to 90 μL of G6 (11.08 μM final concentration) or PAH (13.84 μM final concentration) in NaCl solution. The NaCl concentration depicted is the final concentration of NaCl in the experiment. Polymer concentrations were the same as used for P22@polymer preparation. Samples were left to stand at room temperature overnight, then analyzed on an Agilent Cary 8454 UV–vis Spectrophotometer.

Kraj P., Selivanovitch E., Lee B, & Douglas T. (2021). Polymer Coatings on Virus-like Particle Nanoreactors at Low Ionic Strength—Charge Reversal and Substrate Access. Biomacromolecules, 22(5), 2107-2118.

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Electrochemical Polymer Characterization

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All the chemicals were of reagent grade and used without further purification. All reactions were carried out under an N₂ atmosphere using anhydrous solvents.
¹H and ¹³C NMR spectra were recorded with Bruker AVIII-400 spectrometers, and chemical shifts were measured in δ (ppm) with residual solvent peaks as internal standards. UV–visible (UV/Vis) spectra were measured on an Agilent Technologies Cary 8454 UV-V is spectrophotometer. The mass spectrum was obtained on a TOF-MS spectrometer. The morphology of the electrodeposited polymers on ITO-glass slides was recorded using a field-emission scanning electron microscope (FE-SEM, ULTRA PLUS). Electrochemical polymerizations and electrochemical studies were performed using an Autolab potentiostat (PGSTAT128N, ECO CHEMIE BV, the Netherlands). The XPS spectra were acquired with a PHI 5000 VersaProbe (ULVAC-PHI, Chigasaki, Japan) spectrometer with a 24.7 W micro focused Al kα X-ray source and a take-off angle of photoelectron at 45°. The fluorescence microscopy images were recorded with a Nikon–ECLIPS Ni-E microscope (Nikon Corporation, Tokyo, Japan) and Olympus IX81 fluorescence microscope (Japan).

Ayalew H., Ali S.A., She J.W, & Yu H.H. (2022). Biguanide- and Oligo(Ethylene Glycol)-Functionalized Poly(3,4-Ethylenedioxythiophene): Electroactive, Antimicrobial, and Antifouling Surface Coatings. Frontiers in Chemistry, 10, 955260.

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Characterization of Organometallic Complexes

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NMR spectra were recorded at room temperature using a JEOL ECA-600, ECA-500, or ECA-400 NMR spectrometer. Infrared (IR) spectra were measured using a Thermo Nicolet Avatar FT-IR spectrometer with diamond ATR. UV-vis absorption spectra were recorded in THF, toluene, and MeOH solutions in screw-capped 1 cm quartz cuvettes using an Agilent Cary 8454 UV-vis spectrophotometer. Cyclic voltammetry (CV) measurements were performed with a CH Instruments 602E potentiostat interfaced with a nitrogen glovebox via wire feedthroughs. Samples were dissolved in dichloromethane (CH2Cl2) with 0.1 M TBAPF₆ as a supporting electrolyte. A 3 mm diameter glassy carbon working electrode, a platinum wire counter electrode, and a silver wire pseudo-reference electrode were used. Potentials were referenced to an internal standard of ferrocene. The bulk purity for all complexes is established by elemental analysis, performed by Atlantic Microlab, Inc.

Mu G., Wen Z., Wu J.I, & Teets T.S. (2019). Azo-triazolide bis-cyclometalated Ir(III) complexes via cyclization of 3-cyanodiarylformazanate ligands. Dalton transactions (Cambridge, England : 2003), 49(12), 3775-3785.

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Spectroscopic Characterization of Organic Compounds

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All the biological analytes
and other chemical reagents were purchased from commercial suppliers
(Sigma Aldrich, Tokyo Chemical Industry, Alfa Aesar, and UChem) and
used without further purification. All melting points were recorded
on a micromelting point apparatus and are stated uncorrected. Reactions
were monitored by thin layer chromatography (TLC) with 0.25 mm precoated
silica gel plates (Kieselgel 60F₂₅₄). Flash column chromatography
was performed on silica gel (70–230 mesh) using distilled organic
solvents. Fluorescence spectra were recorded using a Cytation 3 Multi-Mode
Reader and an Agilent Cary Eclipse fluorescence spectrophotometer.
All UV–vis spectra were recorded using an Agilent Cary 8454
UV–vis spectrophotometer. ¹H NMR and ¹³C{¹H} NMR spectra were obtained using a Bruker Advance
III HD 600 MHz spectrometer. Chemical shifts are reported as δ
(ppm) values relative to chloroform (CDCl₃, δ 7.260)
and dimethyl sulfoxide (DMSO-d₆, δ
2.50), and coupling constants are reported in Hz. High-resolution
mass spectroscopy (HRMS) data were obtained using a magnetic sector-electric
sector double focusing mass analyzer at the Korea Basic Science Institute
(KBSI) in electron spray ionization (ESI) mode and measured by time
of flight (TOF) in ESI mode on an SCIEX X500R QTOF.

Lee S., Sung D.B., Lee J.S, & Han M.S. (2020). A Fluorescent Probe for Selective Facile Detection of H2S in Serum Based on an Albumin-Binding Fluorophore and Effective Masking Reagent. ACS Omega, 5(50), 32507-32514.

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AChE Inhibition Activity Assay

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Acetylcholinesterase (AChE) inhibition activity was
measured using Ellman’s method as described by Mathew and Subramanian
(2014) with some modifications.^{53 (link)} In brief,
100 μL of 3 mM of DTNB (5,5-dithiobis[2-nitrobenzoic acid])
dissolved in 50 mM Tris-HCl buffer (pH 8.0) containing 0.1 M NaCl
and 0.02 M MgCl₂, 20 μL of 0.26 U/mL AChE dissolved
in 0.1% BSA (bovine serum albumin) in buffer, 640 μL of buffer,
and 20 μL of the extract were mixed. After incubation for 15
min at 25 °C, absorbance was measured at 412 nm in a Cary 8454
UV–Vis spectrophotometer (Agilent Technologies), which was
treated as the control. Then, the enzymatic reaction was started by
the addition of 15 mM ATCI (acetylthiocholine iodide) dissolved in
water, and the absorbance was read at 412 nm until the reaction completed
(45 min). Galantamine (100 μM) was used as the positive control.
The results were expressed as % inhibition of AChE employing the following
equation: where Abs_control is the absorbance
containing all reagents except ATCI and Abs_sample is the
absorbance of the solution prepared after completing the enzymatic
reaction with ATCI.

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Antiaging Elastase Inhibition Assay

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Elastase
inhibition activity assay based on Azmi et al. (2014) with some modifications
was employed to determine the antiaging capacity of the extracts.^{52 (link)} Briefly, 100 μL of 0.2 mM Tris-HCl buffer
(pH 8.0), 25 μL of 10 mM N-succinyl-Ala-Ala-Ala-p-nitroanilide dissolved in the Tris-HCl buffer, and 50
μL of extract were mixed. After incubation for 15 min at 25
°C, absorbance was measured at 410 nm. Then, 25 μL of 0.3
units/mL elastase was added and incubated for another 15 min at 25
°C and the absorbance was read at 410 nm in a Cary 8454 UV–Vis
spectrophotometer (Agilent Technologies, Palo Alto, CA, USA). Epicatechin
(0.7 mg/mL) was used as a positive control. The results were expressed
as % of elastase inhibition activity employing the following equation: where C is the absorbance
of the extract after incubation with the enzyme, D is the absorbance of the extract after incubation without enzyme, A is the absorbance of the control after incubation with
enzyme and B is the absorbance of the control after
incubation without enzyme.

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