An Investigation of the Antitumor Activity of Micromelum integerrimum (Rutaceae)

Cassady, John M.; Ojima, Nobutoshi; Chang, Ching-jer; McLaughlin, Jerry L.

doi:10.1021/np50003a005

Cited by 54 publications

(39 citation statements)

References 6 publications

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“…The stock solution was kept in a dark bottle and was found to be stable at 4°C for several weeks. Its concentration was determined by measuring the absorbance at 346 nm where the molar absorption coefficient is 1…”

Section: Methodsmentioning

confidence: 99%

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Transient and Steady‐State Kinetics of the Oxidation of Scopoletin by Horseradish Peroxidase Compounds I, II and III in the Presence of NADH

1995

European Journal of Biochemistry

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Scopoletin, a naturally occurring fluorescent component of some plants and a proven plant growth inhibitor, is a known reactant with peroxidase. However, the kinetics of the elementary steps of the reaction have never been investigated, nor has the quantitative effect of interfering substances ever been explored in detail, despite the fact that scopoletin is widely used in a peroxidase assay for H,O,. In this work, we employed both transient-state and steady-state methods to determine the second-order rate constants for the oxidation of scopoletin by the horseradish peroxidase (HRP) intermediate compounds I and 11: (3.7-t0.1)X10h M~' s I and (8.5-+0.5)X1Os M --' s .I at 20°C, pH 6.0 and ionic strength of 0.1 M. We investigated the possible inhibitory effect of NADH on the reaction of scopoletin with HRP and also the effect of scopoletin on the NADH reaction. In the presence of NADH the rate constant for the reaction between HRP-I and scopoletin decreased slightly to (2.820.1)XlO" M I s -' . Thus, although NADH is also a peroxidase substrate, it cannot compete effectively for the oxidized forms of the enzyme. On the other hand, scopoletin stimulates the oxidation of NADH by the HRP/H,O, system, apparently by forming a phenoxyl radical which then oxidizes NADH to NAD' radicals. We present spectral evidence showing that in the aerobic reaction between HRP and NADH at pH 7.0 (without exogenously added HZOJ HRP-I1 is the dominant enzyme intermediate with HRP-I11 also detectable. Addition of scopoletin to the HRP/ NADH system leads to a biphasic reaction in which HRP-I1 and HRP-I11 disappear. The rate constants for both phases are linearly dependent on scopoletin concentration. We attribute the faster phase to the HRP-I1 reaction with scopoletin with a rate constant of (6.2f0.1)X10s M -' s~-' and the slower phase to the HRP-I11 reaction with scopoletin with rate constant (5.0t0.4)X104 M ' s '. Our present work not only provides rate constants for the oxidation of scopoletin by HRP-I, I1 and 111 but also elucidates the interactions that possibly occur physiologically during NADH oxidation in the presence of scopoletin.Keywords: scopoletin; NADH ; horseradish peroxidase; transient-state kinetics; steady-state kinetics. Scopoletin (7-hydroxy-6-methoxy-2H-l-benzopyran-2-one)is a pharmacologically active agent that has been isolated from several plant species [I -31. It contributes to the blue fluorescence of cotton leaves [4, 51 and proves to be the most active of all coumarin derivatives in causing inhibition of root growth 161. Its localization in plant cells and its ability to serve as an aromatic donor molecule suggest that it could be a physiological substrate of the ubiquitous peroxidase enzymes. Scopoletin has been found to be the most potent naturally occurring stimulator of indoleacetic acid oxidase activity 171. Since phenolic compounds are known enhancers of peroxidase-catalyzed oxidations [8], we were interested not only in scopoletin's behavior as a peroxidase substrate but also how it affects the oxidation o...

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Section: Methodsmentioning

confidence: 99%

“…The effect of catalase was explored by incubating the NADH solution first with 20 nM catalase before adding it to HRP. The effect of superoxide dismutase was determined by adding 1 . O pM of the enzyme to HRP before NADH was introduced to the system.…”

Section: Methodsmentioning

confidence: 99%

Transient and Steady‐State Kinetics of the Oxidation of Scopoletin by Horseradish Peroxidase Compounds I, II and III in the Presence of NADH

1995

European Journal of Biochemistry

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“…The HPLC-DAD and UPLC-ESI(+)-HRMS and HRMS/MS profiling of different extracts of M. falcatum revealed useful spectroscopic and spectrometric features which enabled the detection of numerous coumarin derivatives and led to a targeted isolation thereof. In total, eight 7-methoxylated coumarins, among them the three new natural products microfalcrin (1), microcoumaririn (2), micromelosidester (3) and the known ones micromeloside A (4) [8], microminutin (5) [11], micromelin (6) [14], micromarin B (7) [15] and micromarin A (8) [15] were isolated and structurally elucidated. In addition, the inhibition of the NO synthase and the pro-inflammatory agent NF-κB by representative extracts and compounds was explored.…”

Section: Introductionmentioning

confidence: 99%

Rapid Identification of Coumarins from Micromelum falcatum by UPLC-HRMS/MS and Targeted Isolation of Three New Derivatives

et al. 2014

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Micromelum falcatum, a medicinal plant of the Rutaceae family, has been used in the Traditional Chinese Medicine (TCM) mainly against colds and rheumatoid arthritis. Despite its traditional use the association of its constituents with possible anti-inflammatory activity has not been explored. During this study, a rapid UPLC-ESI(+)-HRMS method was developed for the profiling of M. falcatum leave extracts and the targeted isolation of coumarin constituents. Based on chromatographic, spectroscopic and spectrometric features several 7-oxygenated coumarin derivatives were detected. After targeted isolation, eight coumarins, among them three new natural products, namely microfalcrin, microcoumaririn and micromelosidester, were purified using semi-preparative HPLC and unambiguously identified by 1 and 2D NMR. Furthermore, important spectrometric characteristics were revealed based on the HRMS and HRMS/MS spectra of the isolated 7-oxygenated coumarins facilitating their identification in complex mixtures. Finally, the anti-inflammatory properties of the extracts and representative compounds were evaluated OPEN ACCESSMolecules 2014, 19 15043 by measuring the inhibition of the pro-inflammatory mediator NF-κB induction and nitric oxide (NO) production.

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“…The leaves are used traditionally as a febrifuge, the stems as a carminative, and the flowers and fruits as an expectorant and a purgative, respectively [9]. The genus Micromelum species are known to contain 6-and 8-prenylated coumarins [10][11][12]. Micromelum minutum seeds was chosen for current study because the seeds are rich source of coumarins.…”

Section: Introductionmentioning

confidence: 99%

Antihyperglycemic, Antihyperlipidemic and Antiapoptotic Activities of Micromelum minutum Seeds in Diabetic Rats

Koriem¹

2014

J Mol Genet Med

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An Investigation of the Antitumor Activity of Micromelum integerrimum (Rutaceae)

Cited by 54 publications

References 6 publications

Transient and Steady‐State Kinetics of the Oxidation of Scopoletin by Horseradish Peroxidase Compounds I, II and III in the Presence of NADH

Transient and Steady‐State Kinetics of the Oxidation of Scopoletin by Horseradish Peroxidase Compounds I, II and III in the Presence of NADH

Rapid Identification of Coumarins from Micromelum falcatum by UPLC-HRMS/MS and Targeted Isolation of Three New Derivatives

Antihyperglycemic, Antihyperlipidemic and Antiapoptotic Activities of Micromelum minutum Seeds in Diabetic Rats

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