Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.3.1 (alkaline phosphatase)
47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In order to obtain some informations on the nature and relative activity of the phosphatases present in various helminths, biochemical studies have been made in thirteen kinds of worm parasites including the adults and larvae (Fasciola hepatica, Eurytrema pancreaticum, Paramphistomum sp., Taenia solium, Taenia pisiformis, Dipylidium caninum, Diphyllobothrium mansoni, Cysticercus cellulosae, Cysticercus fasciolaris and Sparganum). A comparison based on the analysis of pH-activity curves was made among these helminths. The worm materials were mostly obtained alive from an abattoir and removed from the organs or tissues of the animal hosts naturally infected. Sparganum and Cysticercus cellulosae, however, are collected from the subcutaneous tissue of the patients by surgical removal. The worms thoroughly washed were weighed and transferred with 0.1 M Tris buffer to a chilled glass grinder (Capacity; 15 ml) and homogenized in the cold. The homogenate was centrifuged at 5000 RPM for 30 minutes. The supernatant was pipetted off for determination of the phosphatase activity. Incubation mixtures consisted of 1 ml substrate, 1 ml buffer and 0.5ml extract. The buffers used were Tris (Hydroxymethyl) aminomethane and citric acid monohydrate and the substrate was paranitrophenyl phosphate (1 gm/25 ml). These mixtures were incubated at the temperature of 37 degrees C for 30 minutes in water bath. The absorbance or transferance of mixture was determined colorimetrically by "Spectronic 20 "spectrophotometer at 410 nm against a distilled water blank. The amount of phenol liberated was then calculated from a standard curve using phenol solutions. Controls consisted of unincubated mixtures. The results were deducted from this experiment. The phosphatase activity occurred over all parasitic helminths used in this experiment. In trematodes, pH-activity curves have demonstrated two peaks of phosphatase activity in Fasciola hepatica and Paramphistomum species. However the acid phosphatase activity was predominantly found and the alkaline phosphatase activity was found distinctly to be low in all three species. In Eurytrema pancreaticum, the pH-activity curves displayed two peaks in acid phosphatase activity, one at pH 5.0 and the other pH 9.0. In cestodes, both alkaline and acid phosphatase activity displayed the pH optima 5.0 and 9.0 to 10.0 in the adult tapeworms. However, major activity in the adults is due to the alkaline phosphtases. In contrast to the adults, Cysticercus and sparganum showed the higher activity in acid phosphatases which predominates in the larvae. In all cases of nematodes, the pH optimum for acid phosphatase was 4.0 to 6.0. A preponderance of acid phosphatase activity was shown in the extract of intestine of Ascaris lumbricoides. The aspect that phosphatases are correlated with phosphorylated passage of substances through the cuticle of helminths and may also be involved in carbohydrate metabolism is discussed.
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PMID:[Studies On Phosphatase Activity In Some Parasitic Helminths] 1291 52

Chromatographic behavior of whole type 1 poliovirus and phenol-extracted viral RNA on diethylaminoethyl cellulose columns, as revealed by assay of plaque-forming capacity, indicated that infectious RNA had surface properties markedly different from those of the intact virus. Infectious RNA of type 1 poliovirus and Coxsackie B1 virus was relatively resistant to heat inactivation as compared to intact virus. Kinetics of inactivation at elevated temperatures were multi-hit in character. The structure of infectious enterovirus RNA was investigated by treatment with chemical inactivating agents. Urea and guanidine as hydrogen bond-disrupting agents, and mercaptoethanol and thioglycolate as disulfide bond-disrupting agents, and combinations of these did not destroy RNA infectivity whereas hydrogen bond-disrupting treatment inactivated intact virus rapidly. RNA infectivity was not reduced by chloroform extraction alone, or by octanol extraction alone, but was reduced by chloroform-octanol extraction which failed to depolymerize RNA to an extent detectable by ultracentrifugal analysis. Infectivity of type 1 poliovirus and Coxsackie B1 virus RNA was destroyed in accordance with first order kinetics by very dilute solutions of pancreatic ribonuclease, and by purified snake venom phosphodiesterase, but not at all by bacterial alkaline phosphatase. Inactivation by venom diesterase was not accelerated by prior treatment of RNA with bacterial alkaline phosphatase. These results indicated that infectivity of enteroviral RNA resided in a single stranded structure, that a single break of a phosphodiester bond anywhere along the structure was sufficient to destroy infectivity, and that infectivity did not require a terminal phosphate group. Hydroxylamine, but not other carbonyl reagents, rapidly destroyed infectivity of intact type 1 poliovirus viral RNA without depolymerization of RNA-detectable by behavior in the analytical ultracentrifuge. With S(35)-methionine-labeled poliovirus a very small fraction of radioactivity remained in RNA preparations following phenol extraction. No evidence could be obtained to indicate that infectious enteroviral RNA was composed of subunits. RNA extracted with phenol during the course of infection of HeLa cells with type 1 poliovirus resembled RNA obtained from purified whole virus with respect to heat inactivation, hydroxylamine inactivation, chromatographic separation, susceptibility to protein denaturing agents, and ability to infect productively both naturally susceptible HeLa cells and naturally insusceptible L strain mouse cells. Intracellular production of infectious RNA paralleled intracellular maturation of whole virus and preceded it by a very short interval.
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PMID:Enteroviral ribonucleic acid. II. Biological, physical, and chemical studies. 1371 82

Osteoblastic induction is commonly studied using the colony-forming unit-fibroblastic (CFU-f) assay, in which bone marrow stromal cells (BMC) are grown in a tissue culture environment permissive for osteoblastic differentiation (DMEM containing dexamethasone, ascorbic acid and beta-glycerophosphate). These cells form colonies, which express alkaline phosphatase, and form a collagenous matrix that becomes calcified. However, these same cells originate in the bone marrow where under normal circumstances they do not proliferate or differentiate despite being subjected to many of the same growth factors and hormones present within the tissue culture environment. We show here that phenol red, present within tissue culture medium as a pH indicator, may itself be a factor that permits osteoblastic recruitment. BMC cultured in the presence of the bone anabolic agents PGE2, PGA2, or bFGF, but in the absence of phenol red, failed to respond to these agents in terms of total or osteoblastic colony number. This effect was dose dependent, with low (2.5 mg/l) and high (15-20 mg/l) doses of phenol red being nonpermissive for the stimulatory effects of PGE2 whereas doses of 5-10 mg/l were permissive. Furthermore, the effects observed in the absence of phenol red could not be abrogated by the addition of 17beta-estradiol indicating that these effects cannot be attributed to estrogenic impurities within the phenol red preparation. This indicates that phenol red itself can affect the differentiation of BMC by a mechanism not previously described.
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PMID:Effects of phenol red on CFU-f differentiation and formation. 1456 99

A novel method for the determination of alkaline phosphatase (ALP) isoenzymes in individual fibroblast cells of mouse bone marrow was developed by combining capillary electrophoresis with an on-capillary enzyme-catalyzed reaction and electrochemical detection. In this method, a single an cell, followed by 5.0 x 10(-2) mol/L Na2B4O7- 3.0 x 10(-2) mol/L NaCl (pH 9.8) as cell lysis solution, was injected into the inlet of the separation capillary by electromigration. The cell was lysed by applying a voltage of 2 kV. The ALP isoenzymes in the cell were preseparated at 20 kV for 1 min, and then allowed to react for 30 min with disodium phenyl phosphate as enzyme substrate in the running buffer. ALP converted disodium phenyl phosphate into its product, phenol, at a relatively high reaction rate without consumption, with resultant amplification of the signal on prolonged reaction time, producing an adequate amount of product for final detection. A mass detection limit as low as 3.5 x 10(-21) mol/L (corresponding to 1.5 nU) was achieved. Finally, the two zones of products generated by ALP isoenzymes were detected at the outlet of the capillary by using the end-capillary amperometric detection at a carbon fiber microdisk bundle electrode with a constant potential.
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PMID:Measurement of alkaline phosphatase isoenzymes in individual mouse bone marrow fibroblast cells based on capillary electrophoresis with on-capillary enzyme-catalyzed reaction and electrochemical detection. 1521 85

The alkaline phosphatase test is used as an indicator of adequate pasteurisation of milk and cream. A proprietary fluorimetric technique (Fluorophos) is a sensitive and quantitative method for the determination of alkaline phosphatase (ALP) activity in milk products. Currently, adequate pasteurisation of milk products is regarded as confirmed in samples that contain a residual bovine ALP activity of < or =500 mU/litre. This is equivalent to the statutory acceptable level of 4ug phenol/ml required by the EC analytical method. The purpose of the present study was to assess the effectiveness of pasteurisation of milk and cream produced by on-farm dairies. In a longitudinal study over a four-year period, 4,999 samples of milk and cream were collected from 130 on-farm dairies and from two large commercial dairies in NW England for comparison. Bovine ALP activity of >500 mU/litre was deemed as a failure and was found in 3.5% of whole milk, 2.4% semiskimmed milk, 5.0% of skimmed milk, and 39% of cream samples from on-farm dairies. Bovine ALP activity of >100 and <500 mU/litre was found in 18.4% of whole milk, 9.3% of semi-skimmed milk, 13.2% skimmed milk and 44.5% of cream samples from on-farm dairies. Results with skimmed milk samples showed significantly lower bovine ALP activity than whole milk. All 409 milk and cream samples from two large commercial dairies passed the fluorimetric test at less than 500 mU/litre of bovine ALP, and 99% of these milk and cream samples had bovine ALP activity of less than 100 mU/litre. The presence of residual bovine phosphatase indicates a failure and may be due to either inadequate pasteurisation or post pasteurisation contamination with raw milk. Residual bovine phosphatase was demonstrated in 108/114 (94.7%) of milk samples with a bovine ALP activity greater than 500 mU/litre, i.e. true failures. Of more concern is that residual bovine phosphatase was found in 395/401 (98.5%) of samples that gave bovine ALP activity greater than 100 mU/litre but equal to or less than 500 mU/litre. Residual bovine phosphatase was demonstrated in 37/108 (30.2%) of cream samples with bovine ALP activity greater than 500 mU/litre. Presence of reactivated bovine phosphatase is not an indication of a failure but can mask the presence of residual bovine phosphatase. Reactivated bovine phosphatase was found in 74/106 (69.8%) of cream samples. Our results confirm that the more sensitive fluorimetric method is suitable for testing pasteurised whole milk and semiskimmed milk, but for statutory purposes the acceptable level of residual bovine phosphatase should be <100 mU/litre. Our findings have highlighted a potential problem when testing skimmed milk and cream samples from on-farm dairies. To ensure public safety we need more stringent standards for the ALP test and new methods that will accurately confirm that pasteurisation of these products has been achieved.
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PMID:Assessment of pasteurisation of milk and cream produced by on-farm dairies using a fluorimetric method for alkaline phosphatase activity. 1525 5

Hydroquinone diphosphate (HQDP) was synthesized and compared to phenyl phosphate (PP) and 1-naphthyl phosphate (NP) as a substrate for alkaline phosphatase (AP) under electrochemical immunoassay (EIA) conditions. Voltammetric and amperometric experiments showed that electrochemical oxidation of hydroquinone (HQ), which is the AP hydrolysis product of HQDP, did not produce electrode passivation, even with repeated biosensor use. In contrast, phenol and 1-naphthol, the hydrolysis products of PP and NP, respectively, were shown to be irreversibly oxidized on the electrode surfaces, and produced rapid electrode passivation, resulting in complete loss of electrode signal. When employed as AP substrate in an iridium oxide based EIA, HQDP produced significantly larger amperometric responses (117 microA/cm2) compared to PP (31 microA/cm2) and NP (27 microA/cm2). The results presented in this paper show that HQDP is an attractive alternative to commonly used AP substrates such as NP and PP. The substrate shows excellent hydrolytic stability, produces larger amperometric responses (than PP or NP), and does not produce sensor passivation.
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PMID:Hydroquinone diphosphate: an alkaline phosphatase substrate that does not produce electrode fouling in electrochemical immunoassays. 1530 32

The use of an amperometric graphite-Teflon composite tyrosinase biosensor for the rapid monitoring of alkaline phosphatase (ALP), with no need of an incubation step and using phenyl phosphate as the substrate, is reported. Phenol generated by the action of ALP is monitored at the tyrosinase composite electrode through the electrochemical reduction of the o-quinone produced to catechol, which produces a cycle between the tyrosinase substrate and the electroactive product, giving rise to the amplification of the biosensor response and to the sensitive detection of ALP. The current was measured at -0.10 V 5 min after the addition of ALP. As a compromise between high ALP activity and high sensitivity for the detection of phenol, a pH of 8.5 was chosen. The substrate concentration was also optimized. A linear calibration plot was obtained for ALP between 2.0 x 10(-13) and 2.5 x 10(-11), with a detection limit of 6.7 x 10(-14) M. Different types of milk were analyzed with good results, using an extremely simple and rapid procedure.
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PMID:Rapid and highly sensitive electrochemical determination of alkaline phosphatase using a composite tyrosinase biosensor. 1562 Aug 94

The comparative decomposition of chickpea residue, and chopped and unchopped wheat straw was investigated in pits for 120 days. Microbial biomass, humus, C/N ratio, pH, Electrical conductivity (EC), dehydrogenase, alkaline phosphatase, cellulase, xylanase, total phenol and soluble protein were determined to assess their response to the addition of inorganic nitrogen and mixed fungal inoculum of Aspergillus nidulans, Phanerochaete chrysosporium and Trichoderma viride. The evaluation of physico-chemical parameters (organic matter, organic carbon, N, C/N, pH, EC, microbial biomass) revealed that by supplementing unchopped wheat straw with 1% urea and mixed fungal inoculum, a lowest C/N ratio of 10.7, lowest biomass of 9.54 and highest humus content of 13% can be achieved within 3 months. Germination of Lepidium sativum (cress seeds) showed a germination index >60%, in this treatment. The enzyme assay for dehydrogenase indicated highest microbial activity in uninoculated treatments compared to fungal inoculated counterparts, in the second month sampling (active phase of composting). However, cellulase and xylanase activity showed an upward trend during curing phase of composting. Chickpea residue compost, though resulted in a C/N ratio of 17.3, but its germination index was less than 60%. The rapid quality tests conducted for H2S, NH3, NO3 and starch confirmed the stability and maturity of finished compost prepared from wheat straw through microbial inoculants.
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PMID:Biodegradation study of crop residues as affected by exogenous inorganic nitrogen and fungal inoculants. 1602 2

The first hyperpolarizabilities of p-nitrophenol and p-nitrophenylphosphate have been investigated in vacuum and in neutral aqueous solution by means of time-dependent density functional theory. The calculated excited states and hyperpolarizabilities obtained for these systems and for the molecules of phenol, nitrobenzene, and p-nitroaniline in vacuum match well with the experimental trends. The water solvent has been described by the conductorlike screening model and has been completed by water molecules interacting by hydrogen bonds with the solute. The results show a significant effect of the solvent on the first hyperpolarizability. In particular, the hyperpolarizability of p-nitrophenylphosphate (6.78 x 10(-30) esu) in vacuum is only 1.2 times larger than p-nitrophenol (5.63 x 10(-30) esu), whereas it is almost twice higher in aqueous environment, 12.6 x 10(-30) and 6.5 x 10(-30) esu, respectively. This difference in the nonlinear response in neutral water makes the p-nitrophenylphosphate substrate a suitable probe for measuring the activity of alkaline phosphatase enzymes.
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PMID:Water solvent effect on the first hyperpolarizability of p-nitrophenol and p-nitrophenylphosphate: a time-dependent density functional study. 1668 78

A novel progesterone immunosensor using a colloidal gold-graphite-Teflon-tyrosinase composite biosensor as amperometric transducer is reported. A sequential competitive configuration between the analyte and progesterone labelled with alkaline phosphatase (AP) was used. Phenyl phosphate was employed as the AP-substrate and the enzyme reaction product, phenol, was oxidized by tyrosinase to o-quinone, which is subsequently reduced at -0.1 V at the biocomposite electrode. Variables such as the concentration of phenyl phosphate, the amount of antibody attached to the electrode surface, immersion time in a 2% BSA solution, working pH and incubation times in progesterone and AP conjugate were optimized. A linear calibration graph for progesterone was obtained between 0 and 40 ng mL(-1) with a slope value of -82.3 nA ng(-1) mL, and a detection limit of 0.43 ng mL(-1). The time needed to reach the steady-state current from the addition of phenyl phosphate was 30-40 s. These analytical characteristics improve substantially those reported for other progesterone immunosensors. A lifetime of 14 days with no need to apply any regeneration procedure was also achieved. The usefulness of the immunosensor was evaluated by determining progesterone in milk samples spiked with the analyte at 5.0 and 1.5 ng mL(-1) concentration levels. Following a very simple procedure, involving only sample dilution, mean recoveries (n=7) of 98+/-3% and 99+/-3%, respectively, were obtained.
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PMID:Nanostructured progesterone immunosensor using a tyrosinase-colloidal gold-graphite-Teflon biosensor as amperometric transducer. 1761 44


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