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2,2-Diphenyl-1-picrylhydrazyl radical-scavenging active components from Polygonum multiflorum thunb.

An activity-directed fractionation and purification process was used to identify the antioxidative components of Polygonum multiflorum Thunb. (PM). Dried root of PM was extracted with 95% ethanol and then separated into water, ethyl acetate, and hexane fractions. Among these only the ethyl acetate phase showed strong antioxidant activity by the 2,2-diphenyl-1-picrylhydrazyl (DPPH) test when compared with water and hexane phases. The ethyl acetate fraction was then subjected to separation and purification using silica gel column chromatography and Sephadex LH-20 chromatography. Three compounds showing strong antioxidant activity were identified by spectral methods ((1)H NMR, (13)C NMR, and MS) and by comparison with authentic samples to be gallic acid, catechin, and 2,3,5, 4'-tetrahydroxystilbene 2-O-beta-D-glucopyranoside.

Antioxidants↗

The cytochrome P-450 monooxygenase system of rabbit lung enzyme components, activities, and induction in the nonciliated bronchiolar epithelial (Clara) cell, alveolar type II cell, and alveolar macrophage.

Enzyme components and activities of the cytochrome P-450 monooxygenase system in microsomal preparations from the Clara cell, alveolar type II cell, and alveolar macrophage fractions isolated from lungs of untreated rabbits and rabbits treated with 2,3,7,8-tetrachlorodibenzo-p-dioxin were examined. Results are compared to those obtained with microsomal preparations from whole lung. Concentrations of cytochrome P-450 isozymes 2 and 5 and NADPH-cytochrome P-450 reductase activities were higher in preparations from Clara cell fractions than in preparations from type II cell fractions or whole lung. For the most part, however, differences among these preparations were 2-fold or less. Microsomal preparations from the macrophage fraction contained low or undetectable levels of cytochrome P-450 isozymes but relatively high levels of cytochrome P-450 reductase activity. The concentration of cytochrome P-450 isozyme 6, in contrast to those of isozymes 2 and 5, was found to be highest in microsomal preparations from whole lung. Treatment of rabbits with 2,3,7,8-tetrachlorodibenzo-p-dioxin increased the concentrations of isozyme 6 in preparations from the Clara and type II cell fractions and from whole lung about 20-fold. In contrast, the content of isozyme 6 in preparations from the macrophage fraction increased greater than 90-fold. In all cases, induction of isozyme 6 resulted in substantial increases in the O-deethylation of 7-ethoxyresorufin and only minor increases in the hydroxylation of benzo(a)pyrene. Activities per unit of isozyme 6, following induction, were similar in all preparations, and we estimate that less than 20% of the potential activity of isozyme 6 is expressed with benzo(a)pyrene and greater than 40% with 7-ethoxyresorufin. These similarities exist in spite of significant differences among the preparations from different fractions in the ratios of isozyme 6 to NADPH-cytochrome P-450 reductase.

Animals↗

High-performance liquid chromatographic method for simultaneous determination of hawthorn active components in rat plasma.

A simple HPLC method with photodiode-array (PDA) ultraviolet detection was developed for the simultaneous determination of four active polyphenol components of hawthorn (Crataegus), chlorogenic acid, epicatechin, hyperoside and isoquercitrin, in rat plasma. Following extraction from the plasma samples with ethyl acetate-methanol (2:1, v/v), these four compounds were successfully separated using a C18 column with a gradient elution of 5 and 25% acetonitrile in 25 mM phosphate buffer (pH 2.4). The flow-rate was set at 1 ml/min and the eluent was detected at 325 nm for chlorogenic acid, 278 nm for epicatechin, and 360 nm for both hyperoside and isoquercitrin. Narignin (0.82 microg) was used as the internal standard and was detected at 278 nm. The method is linear over the studied range of 0.16-40, 0.63-160, 0.13-32 and 0.13-30 microg/ml for chlorogenic acid, epicatechin, hyperoside and isoquercitrin, respectively. The correlation coefficient for each analyte was greater than 0.995. The intra-day and inter-day precision of the analysis was better than 4 and 7%, respectively. The extraction recoveries at low to high concentration were greater than 85% for both epicatechin and chlorogenic acid, and greater than 94% for both hyperoside and isoquercitrin. The detection limits were 0.04, 0.20, 0.03 and 0.03 microg/ml for chlorogenic acid, epicatechin, hyperoside and isoquercitrin. The developed method was used to analyze the plasma concentrations of the four analytes after the intravenous administration of hawthorn polyphenol extract to rats.

Animals↗

Importance of the active component of the detrusor muscle in bladder compliance.

An appropriate experimental study was designed and carried out in mongrel dogs, in order to evaluate the active muscle and cholinergic participation on bladder compliance. The filling bladder pressure was measured at 100 ml of bladder volume, in 50 dogs distributed into five groups of 10 dogs each: (1) control group, (2) group which received atropine (cholinergic antagonist), (3) group which received verapamil (extracellular Ca2+ blocker), (4) group which received nitroprusside (intracellular Ca2+ antagonist), and (5) group which received EGTA (Ca(2+)-chelating agent). Furthermore, the following was demonstrated. (1) The greatest decrease of the bladder filling pressure was observed in the group which was treated with EGTA. (2) A significant decrease of the bladder filling pressure was also seen in the group which was given nitroprusside. (3) The decrease of bladder filling pressure in the verapamil group tended towards statistical significance. (4) There was no decrease in the atropine group. Thus, it was concluded that the active muscle component has an important role in the bladder compliance (through the intracellular calcium fraction), and the cholinergic component does not participate in the bladder compliance.

Animals↗

Novel opioid peptides derived from casein (beta-casomorphins). II. Structure of active components from bovine casein peptone.

Material with opioid activity had been isolated from an enzymatic casein digest. It was shown to contain a pure heptapeptide with the sequence Tyr-Pro-Phe-Pro-Gly-Pro-Ile. The identity between the opioid principle and the peptide was proven by the fact that chemical reagents or enzymes effecting one would effect the other. After carboxypeptidase Y digestion a pentapeptide, Tyr-Pro-Phe-Pro-Gly, could be isolated; this peptide showed a higher opioid activity than the heptapeptide. The opioid peptides were highly resistant towards proteolysis, even by pronase. The sequence of the hepatapeptide identified it as a fragment of bovine beta-casein. Therefore it was named beta-casomorphin.

Amino Acid Sequence↗

DNA breaking activity and mutagenicity of soy sauce: characterization of the active components and identification of 4-hydroxy-5-methyl-3(2H)-furanone.

Soy sauce is a seasoning consumed widely in Southeast Asia. When supercoiled DNA was incubated with soy sauce at pH7.4 and 37 degrees C, extensive breaking of DNA single-strands was caused. It was found that the breaking activity was due to multiple components with different molecular weight and polarity. One of the components with the breaking activity was purified successively by extraction with ethyl acetate, thin-layer chromatography and high performance liquid chromatography, and identified as 4-hydroxy-5-methyl-3(2H)-furanone (HMF), one of the fragrant components in soy sauce. Formation of this component was found due to Maillard reaction of pentoses/amino acids. HMF was readily degraded into the compound with an endiol-ketol structure and reducing activity. Using 5,5-dimethyl-1-pyrroline N-oxide (DMPO) in an electron spin resonance-spin trapping technique, generation of hydroxyl radical in an aqueous solution of HMF was confirmed. While DNA breaking by soy sauce was little inhibited by the scavengers of active oxygen radicals, the breaking by HMF was effectively inhibited by superoxide dismutase, catalase, hydroxyl radical scavengers, spin-trapping agents, thiol compounds and metal chelating agents. Hence, DNA breaking activity of HMF was found due to generation of active oxygen radicals. HMF was found mutagenic to Salmonella bacteria without metabolic activation, probably due to generation of active oxygen radicals.

DNA↗

Lesions in many different spindle components activate the spindle checkpoint in the budding yeast Saccharomyces cerevisiae.

The spindle checkpoint arrests cells in mitosis in response to defects in the assembly of the mitotic spindle or errors in chromosome alignment. We determined which spindle defects the checkpoint can detect by examining the interaction of mutations that compromise the checkpoint (mad1, mad2, and mad3) with those that damage various structural components of the spindle. Defects in microtubule polymerization, spindle pole body duplication, microtubule motors, and kinetochore components all activate the MAD-dependent checkpoint. In contrast, the cell cycle arrest caused by mutations that induce DNA damage (cdc13), inactivate the cyclin proteolysis machinery (cdc16 and cdc23), or arrest cells in anaphase (cdc15) is independent of the spindle checkpoint.

Cell Division↗

Large scale isolation of functionally active components of the human complement system.

In the present work a scheme is presented for the isolation of multiple components of human complement in a functionally and biochemically pure state and with full hemolytic activity. These preparative procedures allow high recovery of milligram and gram quantities of particular complement components from a large pool (2-11 liters) of fresh EDTA plasma in no more than four chromatographic steps. Many components (C3bINA, C5, C3, C1EI, C4, and C9) are recovered functionally pure or highly purified following the first chromatographic step employing DEAE-Sephacel and may be utilized as reagents with no further purification. Prior to anion exchange, individual units of plasma are treated with inhibitors of complement activation and serum proteases, the pooled plasma is fractionated with polyethylene glycol, depleted of plasminogen on Sepharose-lysine, and rapidly ultrafiltered to low ionic strength and high protein concentration. The high degree of resolution of the components on DEAE-Sephacel subsequently obtained is demonstrated by the functional recovery and purification in a representative experiment as indicated (in their order of elution) for the following proteins: C3bINA (24%, 18-fold), C2 (74%, 12-fold), C7 (87%, 14-fold), factor B (55%, 8.7-fold),, C8 (50%, 16-fold), C6 (82%, 25-fold), beta 1H (39%, 12-fold), C5 (62%, 111-fold), C3 (99%, 64-fold), C1EI (42%, 135-fold), C9 (80%, 297-fold), and c4 (78%, 164-fold). Other components separated by these procedures include C1q and C4 binding protein. Additional steps described, which demonstrate the utility and effectiveness of this preparative scheme, have allowed isolation of C3, C5, and C7 as pure components with full hemolytic activity as judged by functional, immunochemical, and physicochemical criteria. C8, also isolated as a homogeneous protein, was recovered with partial hemolytic activity. All these components were recovered in high yield and in the purification as indicated: C3 (61%, 103-fold), C5 (24% 1350-fold), C7 (19%, 2260-fold), and C8 (32%, 547-fold). Complement components C6, beta 1H, factor B, and C2 in addition to C3bINA, C1EI, C4, and C9 are recovered partially purified with good activity and are amenable to further purification.

Complement C3↗

The activating component of the anaerobic ribonucleotide reductase from Escherichia coli. An iron-sulfur center with only three cysteines.

Class III anaerobic ribonucleotide reductase small component, named protein beta, contains a (4Fe-4S) center. Its function is to mediate electron transfer from reduced flavodoxin to S-adenosylmethionine, required for the introduction of a glycyl radical in the large component, named protein alpha, which then becomes active for the reduction of ribonucleotides. By site-directed mutagenesis we demonstrate that the three cysteines of the conserved CXXXCXXC sequence are involved in iron chelation. Such a sequence is also present in the activase of the pyruvate formate-lyase and in the biotin synthase, both carrying an iron-sulfur center involved in reductive activation of S-adenosylmethionine. Even though they are able to bind iron in the (4Fe-4S) form, as shown by Mössbauer spectroscopy, the corresponding Cys to Ala mutants are catalytically inactive. Mutation of the two other cysteines of the protein did not result in inactivation. We thus conclude that the (4Fe-4S) cluster has, in the wild type protein, only three cysteine ligands and a fourth still unidentified ligand.

Amino Acid Sequence↗

[Modification of innate immunity in humans by active components of shark liver oil].

See fish oils affect different systemic reactions innate immunity including. Innate immunity is responsible for immediate pathogen recognition and inactivation. Innate immunity decides also on the type of required immunity development. In the presented paper we have proved that supportive treatment with shark oil components normalize complement level, natural killer cells activity and reactive oxygen intermediates production by peripheral blood leukocytes of peoples suffering from active form rheumatoid arthritis.

Animals↗

Extra activation component of calcium release in frog muscle fibres.

In addition to activating more Ca(2+) release sites via voltage sensors in the t-tubular membranes, it has been proposed that more depolarised voltages enhance activation of Ca(2+) release channels via a voltage-dependent increase in Ca-induced Ca(2+) release (CICR). To test this, release permeability signals in response to voltage-clamp pulses to two voltages, -60 and -45 mV, were compared when Delta[Ca(2+)] was decreased in two kinds of experiments. (1) Addition of 8 mM of the fast Ca(2+) buffer BAPTA to the internal solution decreased release permeability at -45 mV by > 2-fold and did not significantly affect Ca(2+) release at -60 mV. Although some of this decrease may have been due to a decrease in voltage activation at -45 mV - as assessed from measurements of intramembranous charge movement - the results do tend to support a Ca-dependent enhancement with greater depolarisations. (2) Decreasing SR (sarcoplasmic reticulum) Ca content ([Ca(SR)]) should decrease the Ca(2+) flux through an open channel and thereby Delta[Ca(2+)]. Decreasing [Ca(SR)] from > 1000 microM (the physiological range) to < 200 microM decreased release permeability at -45 mV relative to that at -60 mV by > 6-fold, an effect shown to be reversible and not attributable to a decrease in voltage activation at -45 mV. These results indicate a Ca-dependent triggering of Ca(2+) release at more depolarised voltages in addition to that expected by voltage control alone. The enhanced release probably involves CICR and appears to involve another positive feedback mechanism in which Ca(2+) release speeds up the activation of voltage sensors.

Animals↗

External perturbation of the trunk in standing humans differentially activates components of the medial back muscles.

During voluntary arm movements, the medial back muscles are differentially active. It is not known whether differential activity also occurs when the trunk is perturbed unpredictably, when the earliest responses are initiated by short-latency spinal mechanisms rather than voluntary commands. To assess this, in unpredictable and self-initiated conditions, a weight was dropped into a bucket that was held by the standing subject (n = 7). EMG activity was recorded from the deep (Deep MF), superficial (Sup MF) and lateral (Lat MF) lumbar multifidus, the thoracic erector spinae (ES) and the biceps brachii. With unpredictable perturbations, EMG activity was first noted in the biceps brachii, then the thoracic ES, followed synchronously in the components of the multifidus. During self-initiated perturbations, background EMG in the Deep MF increased two- to threefold, and the latency of the loading response decreased in six out of the seven subjects. In Sup MF and Lat MF, this increase in background EMG was not observed, and the latency of the loading response was increased. Short-latency reflex mechanisms do not cause differential action of the medial back muscles when the trunk is loaded. However, during voluntary tasks the central nervous system exerts a 'tuned response', which involves discrete activity in the deep and superficial components of the medial lumbar muscles in a way that varies according to the biomechanical action of the muscle component.

Adult↗

Isolation of antigenically active components from leptospiral serovar-specific lipopolysaccharide antigen by alkaline treatment.

The serovar-specific main antigen (TM antigen) of Leptospira interrogans serovar canicola, which as lipopolysaccharide properties, was treated with 0.1 N sodium hydroxide. This treatment degraded the antigen into two major antigenic components, one of high and one of low molecular weight. The component with the lower molecular weight (approximately 4,000 daltons) consisted mainly of carbohydrates, having lost almost all of the fatty acid and protein components of the original antigen. Although the substance lacked immunoprecipitable activity, it continued to show serovar-specific inhibitory potency in a radioimmunoassay system as well as in a microscopic immunoagglutination reaction of the organisms. The antigenic activity of the compound was also reduced by periodate oxidation as was that of the TM antigen. A component with the same chemical and physicochemical properties was also produced by alkaline treatment from a different serotype TM antigen (serovar kremastos Kyoto), but it showed no antigenic activity.

Bacterial Proteins↗

2,2'-Diphenyl-1-picrylhydrazyl radical-scavenging active components from adlay (Coix lachryma-jobi L. var. ma-yuen Stapf) hulls.

An activity-directed fractionation and purification process was used to identify the antioxidative components of adlay hulls. Hulls of adlay (Coix lachryma-jobi L. var. ma-yuen Stapf) were extracted with methanol and then separated into water, 1-butanol, ethyl acetate, and hexane fractions. The 1-butanol-soluble fraction exhibited greater capacity to scavenge 2,2'-diphenyl-1-picrylhydrazyl (DPPH) radicals when compared with fractions soluble in water, ethyl acetate, and hexane phases. The 1-butanol fraction was then subjected to separation and purification using Diaion HP-20 chromatography, silica gel chromatography, and HPLC. Six compounds showing strong antioxidant activity were identified by spectroscopic methods ((1)H NMR, (13)C NMR, IR, and MS) and by comparison with authentic samples to be coniferyl alcohol (1), syringic acid (2), ferulic acid (3), syringaresinol (4), 4-ketopinoresinol (5), and a new lignan, mayuenolide (6).

Antioxidants↗

Component activities in the autoregressive activity of physiological systems.

The higher-order autoregressive (AR) activity of a physiological system (Sato 1975a,b; Sato et al., 1977) was decomposed into first- and second-order activities. The former time-pattern displays a fast rise and an exponential decay, while the latter exhibits a damped sine wave. Application of this component analysis to EEG showed the first-order activity to coincide with the nonoscillatory delta component. The frequency distribution of the natural, damped and resonance frequencies of the second-order activities was found to show one, two or three modes in each frequency range of the delta, theta and beta rhythms in ninety normal adult EEGs. Hence, the second-order activities agree with the oscillatory delta, theta, alpha and beta rhythms. Some changes were induced not only in the frequency of theta and beta rhythms but also in that of alpha rhythm by over-breathing for about three minutes.

Electroencephalography↗

Reliability of late positive component activity (P3) in healthy elderly adults.

Auditory oddball paradigms (20% targets) were used to elicit components of the event-related brain potential in healthy, late middle-aged and elderly adults (N = 32) in two separate sessions approximately 4.5 months apart. Within each session data were acquired under two conditions: (a) 250 Hz standard, 500 Hz target; and, (b) 1000 Hz standard, 2000 Hz target. Measures of P3 latency and amplitude showed moderate to high reliability (.55 to .89) over the 4.5-month interval under both stimulus conditions using data acquired from the midline frontal (Fz), central (Cz), and parietal (Pz) sites. In the second session, latencies were prolonged in the 500 Hz target tone condition, and amplitudes tended to be reduced in the 2000 Hz target tone condition. These results suggest that P3 measures in studies of older adults are reliable over time, and the findings are discussed in conjunction with prior P3 reliability studies of younger adults.

Aged↗

Binding of active components of compound 48/80 to rat peritoneal mast cells.

The binding characteristics of compound 48/80 were examined using rat mast cells and fractionated 14C-labeled compound 48/80 components at 4 degrees C in vitro where no degranulation of the cells occurred. The binding potencies of these components in the presence of Ca2+ generally paralleled their histamine releasing activities, except in the case of fractions G (decamer) and H (nonamer), both Ca2+-independent releasers, for the binding of which Ca2+ was inhibitory. Scatchard analyses and displacement studies indicated that the mast cells had two types of binding sites with high and low affinities for fractions D (tridecamer, Ca2+-dependent releaser, Kd = 3.41 X 10(-8) M and 3.35 X 10(-6) M) and H (Ca2+-independent releaser, Kd = 1.11 X 10(-7) M and 9.11 X 10(-6) M), respectively. These sites partially overlapped each other, and also the fraction D site partially overlapped the IgE site and the fraction H site overlapped the neurotensin or substance P site.

Animals↗