Acquity photodiode array detector

Manufactured by Waters Corporation

Sourced in United States

The Acquity photodiode array detector is a high-performance liquid chromatography (HPLC) detection instrument manufactured by Waters Corporation. It is designed to provide sensitive and reliable UV-Vis spectroscopic detection of analytes separated by HPLC. The detector utilizes a photodiode array to simultaneously monitor a wide range of wavelengths, enabling the collection of comprehensive spectral data for each eluting peak.

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7 protocols using acquity photodiode array detector

UPLC Analysis of Flavonoids

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For the separation, we used a UPLC system with a pump and ACQUITY UPLC (Waters, Milford, MA, USA), along with an ACQUITY photodiode array detector (Waters). We used the Empower Data System (Waters) to record the detector output. A Waters ACQUITY BEH C₁₈ column (1.7 μm, 2.1 mm × 100 mm) was used for the analysis. The column temperature was maintained at 25°C. The mobile phase was composed of acetonitrile with 0.1% formic acid and water, and the mixture was injected with sample (2 μL) at a fluid velocity of 0.4 mL/min. For the photodiode array detector analysis, acacetin and luteolin were analyzed at 330 nm and apigenin was analyzed at 345 nm.

Jung J.Y., Lee C.W., Park S.M., Jegal K.H., Kim J.K., Park C.A., Cho I.J., Jung D.H., An W.G., Ku S.K., Zhao R, & Kim S.C. (2017). Activation of AMPK by Buddleja officinalis Maxim. Flower Extract Contributes to Protecting Hepatocytes from Oxidative Stress. Evidence-based Complementary and Alternative Medicine : eCAM, 2017, 9253462.

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Comparative Metabolomic Analysis of Fungal Mutants

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The WT and ΔpksAC mutant were grown on PDA for 14 days. Whole-agar chunks with fungal colonies were excised, soaked in 10 ml of ethyl acetate (EtOAc) in 50-ml Falcon tubes, and incubated at 4°C overnight. An aliquot of 200 μl of the extracts was transferred to a microcentrifuge tube, evaporated to dryness, reconstituted with 30 μl of MeOH, and subjected to LC-MS analyses. Chromatographic separation was achieved using an Acquity ultraperformance liquid chromatography (UPLC) system (Waters Corp., Milford, MA, USA), as previously described (50 (link)). MS analysis was performed on an inline Synapt G2-S high-definition mass spectrometry (HDMS; Waters Corp.) time of flight mass spectrometer using the positive-ion electrospray mode for data collection. The UV data were acquired using a wavelength of 210 to 400 nm, resolution of 1.2 nm, and 20 points/s with an Acquity photodiode array detector (Waters Corp.).

Kim W., Lichtenzveig J., Syme R.A., Williams A.H., Peever T.L, & Chen W. (2019). Identification of a Polyketide Synthase Gene Responsible for Ascochitine Biosynthesis in Ascochyta fabae and Its Abrogation in Sister Taxa. mSphere, 4(5), e00622-19.

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Purification and Characterization of Compounds

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Tetrahydrofuran was distilled from sodium benzophenone. All other solvents and chemical reagents were obtained from commercial sources and used directly without further purification. Glassware was oven-dried before use. All reactions were performed under an argon atmosphere. TLC was performed on silica gel 60 GF254 and monitored under UV light or visualized using phosphomolybdic acid reagent. Flash chromatography was performed on 230–400 mesh silica gel (Fisher Scientific). Melting points were recorded on an MPA100 Automated Melting Point Apparatus. NMR spectra were obtained on a Bruker Ascend 400 (Billerica, MA) spectrometer or a Varian Inova-500 spectrometer (Agilent Technologies, Santa Clara, CA). HR-MS were obtained on Waters Acquity UPLC linked to a Waters Acquity Photodiode Array Detector and Waters qTof mass detector. All compounds reported herein with biological data had purities ≥95% as determined by HPLC. The purity of associated compounds was verified by the HPLC study performed on BEH C18 (2.1 × 50 mm, 1.7 μm) column using a mixture of solvent acetonitrile/water (with 0.1% formic acid) at a flow rate of 0.3 mL/min and monitoring by UV absorption at the appropriate wavelength. Chemical shifts are given in ppm with tetramethylsilane (TMS) as an internal reference. All coupling constants (J) are given in hertz (Hz).

Wang Q., Arnst K.E., Wang Y., Kumar G., Ma D., Chen H., Wu Z., Yang J., White S.W., Miller D.D, & Li W. (2018). Structural Modification of the 3,4,5-Trimethoxyphenyl Moiety in the Tubulin Inhibitor VERU-111 Leads to Improved Antiproliferative Activities. Journal of medicinal chemistry, 61(17), 7877-7891.

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Sterol and Carotenoid Analysis in Samples

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According to procedures that were described by Stuper-Szablewska et al. [22 (link)], sterol content was determined. Sterol analysis was done using an Aquity H class UPLC system equipped with a Waters Acquity photodiode array detector (Waters, Milford, MA, USA). Chromatographic separation was achieved on an Acquity UPLC^® BEH C18 column (100 mm × 2.1 mm × 1.7 μm) (Waters, Ireland). The elution was isocratically performed using a mobile phase of: A, acetonitrile 10%; B, methanol 85%; C, water 5%, and flow of 0.5 mL/min. The injection volume was 10 µL. Sterol concentrations were measured using an external standard at wavelengths λ = 210 nm (desmosterol, cholesterol, lanosterol, stigmasterol, β-sitosterol) and λ = 282 nm (ergosterol). Compounds were identified by comparing the retention times of the investigated peak and the reference, as well as by adding a particular quantity of the standard to the tested sample and repeating the tests. The detection limit was 1 mg/kg. Carotenoids were evaluated by utilizing Acquity ultra-high performance liquid chromatography (Waters, Milford, MA, USA) as described by Kurasiak-Popowska et al. [23 (link)].

Alharthi S.S., Badr A.N., Gromadzka K., Stuper-Szablewska K., Abdel-Razek A.G, & Selim K. (2021). Bioactive Molecules of Mandarin Seed Oils Diminish Mycotoxin and the Existence of Fungi. Molecules, 26(23), 7130.

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Synthesis and Characterization of Novel Compounds

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All solvents and chemical reagents were purchased from commercial sources and used directly without further purification. Glassware was oven-dried before use. All reactions were performed under an argon atmosphere. Analytical thin-layer chromatography was performed on silica gel 60 GF254 and monitored under UV light. Flash chromatography was performed on 230–400 mesh silica gel (Fisher Scientific). NMR spectra were obtained on a Bruker Ascend 400 (Billerica, MA) spectrometer. The purity of all final compounds (as stereoisomers not separable on the UPLC column used) was ≥95% as determined by UPLC and further confirmed by proton NMR (details given in the Supporting Information). High-resolution mass spectrometry data (HRMS) were obtained on a Waters Acquity UPLC linked to a Waters Acquity Photodiode Array Detector and a Waters qTof mass detector. UPLC analyses were performed using a BEH C18 (2.1 × 50 mm, 1.7 μm) column and a mixture of solvent methanol/water at a flow rate of 0.3 mL/min, monitored by UV absorption at the appropriate wavelength. Chemical shifts are given in ppm. Tetramethylsilane is used as an internal reference for NMR spectra taken in chloroform-d. All coupling constants (J) are given in hertz (Hz).

Wu Z., Gu L., Zhang S., Liu T., Lukka P.B., Meibohm B., Bollinger J.C., Zhou M, & Li W. (2021). Discovery of N-(3,4-Dimethylphenyl)-4-(4-isobutyrylphenyl)-2,3,3a,4,5,9b-hexahydrofuro[3,2-c]quinoline-8-sulfonamide as a Potent Dual MDM2/XIAP Inhibitor. Journal of medicinal chemistry, 64(4), 1930-1950.

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Chromatographic Evaluation of UPLC Columns

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All chromatographic evaluations were performed using ACQUITY UPLC Classic, H‐Class, or I‐Class instruments equipped with ACQUITY photodiode array detectors (Waters, Milford, MA). ACQUITY UPLC BEH C₁₈, CSH C₁₈, HSS T3, and Atlantis PREMIER BEH C₁₈ AX columns (1.7 or 1.8, 2.5 and 5 μm, 2.1 × 50 mm) were obtained from Waters (Milford, MA).

Walter T.H., Alden B.A., Field J.A., Lawrence N.L., Osterman D.L., Patel A.V, & DeLoffi M.A. (2021). Characterization of a highly stable mixed‐mode reversed‐phase/weak anion‐exchange stationary phase based on hybrid organic/inorganic particles. Journal of Separation Science, 44(5), 1005-1014.

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UPLC Chromatographic Evaluation Methods

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All chromatographic evaluations were performed using ACQUITY UPLC Classic, H‐Class, I‐Class, or Premier Systems equipped with ACQUITY photodiode array detectors (Waters Corporation, Milford, MA, USA). ACQUITY UPLC BEH Amide, BEH HILIC, and Atlantis Premier BEH Z‐HILIC columns (1.7 μm, 2.1 × 50 mm) were obtained from Waters (Milford, MA, USA).

Walter T.H., Alden B.A., Berthelette K., Field J.A., Lawrence N.L., McLaughlin J, & Patel A.V. (2022). Characterization of a highly stable zwitterionic hydrophilic interaction chromatography stationary phase based on hybrid organic–inorganic particles. Journal of Separation Science, 45(8), 1389-1399.

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