The Source of Synovial Fluid Alkaline Phosphatase.

Dabich, Danica; Neuhaus, Otto W.

doi:10.3181/00379727-123-31548

Cited by 9 publications

(5 citation statements)

References 1 publication

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“…Our best gtycoprotein preparations show a specific activity of 1920 U rag_ -1 of protein, using the enzymatic unit reported by Dabich and Neuhaus [14]. These investigators have purified a phosphatase from bovine synovial fluid 2286-fold, obtaining a specific activity of 16 000 U rag-1 of protein.…”

Section: Resultsmentioning

confidence: 58%

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Alkaline phosphatase activity associated to a calcium binding glycoprotein from calf scapula cartilage

Vittur

Bernard

1973

FEBS Letters

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Section: Resultsmentioning

confidence: 58%

“…Thus from our graph a good activity of alkaline phosphatase appears possible, not considering of course the influence of the type of substrate and of its concentration. From a Lineweaver plot we have calculated a K M for p-nitrophenylphosphate of 5.9 × 10-4M, a value consistant with that of other phosphatases [14].…”

Section: Resultsmentioning

confidence: 82%

Alkaline phosphatase activity associated to a calcium binding glycoprotein from calf scapula cartilage

Vittur

Bernard

1973

FEBS Letters

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“…We found that the catalytic activity of alkaline phosphatases from the three sources reached a maximum at pH 9.8-10.0 (at these pH values, the functional groups of enzyme active centers responsible for immobilization and substrate binding underwent ionization [19]) and depended on the nature and concentration of the buffer solution ( Fig. 1) in which the enzymatic hydrolysis of p-NPP was performed.…”

Section: Resultsmentioning

confidence: 98%

“…It is well known [10,11,35] that K, Na, Ca, Fe, Cu, Pb, Hg, chloride, and sulfate ions are constituents of urine, along with magnesium ions. However, the individual interfering effects of these ions on the enzyme were precluded under experimental conditions because we found in preliminary experiments that they (in amounts that usually occur in urine [19]) have no effect on the catalytic activity of alkaline phosphatase from chicken intestine.…”

Section: Determination Of Magnesium In Aqueous Solutionsmentioning

confidence: 97%

An enzymatic method for determining magnesium

2005

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Magnesium deficiency in the human body (hypomagnesemia) is a phenomenon typical of almost all countries of the present-day world, including the industrialized countries. Hypomagnesemia is most often due to low dietary magnesium; however, stress, high levels of physical activity, hypodynamia, alcohol abuse, hormonal contraception, etc., may also be responsible for the development of magnesium deficiency [1]. To diagnose the states of magnesium deficiency, the concentration of magnesium in human blood plasma and cells is usually determined by atomic emission spectrometry (AES) with flame ionization [2], inductively coupled plasma AES [3], atomic absorption spectrometry (AAS) with flame [4] or electrothermal [5] atomization, and spectrophotometry with the use of organic reagents (Magon is most frequently used) [6].Recent medical studies have demonstrated that the possibility of magnesium deficiency cannot also be excluded at a normal concentration (16-30 µ g/mL) in blood plasma and cells [7][8][9]. The diagnosis of magnesium depletion (particularly in diabetic patients or postoperative heart patients) can be reliably performed only with the use of a magnesium loading test, i.e., by administering magnesium-containing drugs to patients [8,9]. Substitution therapy is primarily empirical in the cases of latent magnesium deficiency or the impossibility of determining the total magnesium content of the human body.Thus, the analysis of a single biosubstrate (blood) is insufficient for the correct evaluation of the magnesium content of the human body and for the correction of its deficiency with the use of an appropriate magnesiumcontaining drug. The determination of magnesium in daily urine gives additional information on the level of patient's health and provides an opportunity to develop an appropriate course of treatment. The normal magnesium content of urine from adult humans is 100-300 mg/day [10,11]. Under the conditions of magnesium deficiency, the daily excretion of magnesium can be lower than 25 mg/day (0.5 mmol/day).In clinical laboratories, urine magnesium is determined by chelatometric titration [11], AAS with flame and electrothermal atomization [12,13], and spectrophotometry with the use of Xylidyl Blue [14,15] or Magon [8]. Currently available procedures for the determination of magnesium in urine are accurate and inexpensive but insufficiently sensitive (chelatometry), simple but insufficiently accurate (spectrophotometry), or require the use of expensive instrumentation and the removal of spectral interferences (AAS). Moreover, procedures for the simple and reliable determination of magnesium in urine at a low concentration level (ng/mL) were not described in the literature. Consequently, the development of highly sensitive and selective (in particular, enzymatic) procedures for the determination of magnesium is of particular current interest for the accurate evaluation of the magnesium content of biological systems and the subsequent therapy. The use of enzymes of different origins, such as alkaline phosp...

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“…Surprisingly, the enzyme was insensitive to diisopropylfluorophosphate [up to 10 mM (N. H. Georgopapadakou, unpublished)], an organophosphorus reagent that generally inhibits serine hydrolases [alkaline phosphatase being a notable exception (21,22)]. It is also insensitive to phenylmethylsulfonyl fluoride, another, although less potent, inhibitor of several serine hydrolases (23).…”

Section: And 9)mentioning

confidence: 99%

Isolation of the penicillin-binding peptide from D-alanine carboxypeptidase of Bacillus subtilis.

Georgopapadakou¹,

Hammarström²,

Strominger³

1977

Proc. Natl. Acad. Sci. U.S.A.

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Preparation of Labeled CPase. CPase was isolated by affinity chromatography (4) from B. subtilis (strain Porton) membranes obtained by lysozyme DNase treatment (11). In a typical experiment, 10 mg of enzyme was obtained from 300 g of cells. The enzyme in 5 ml of 0.05 M Tris-HCI buffer (pH 7.5) containing 1% (vol/vol) Triton X-100, 0.5 M NaCl and 1 mM 2-mercaptoethanol] was incubated with labeled penicillin G ([8-14C]penicillin G,25 ,uCi, or [35S]penicillin G, 0.8,Ci) for 10 min at 25°. The incubation was stopped by adding 4 volumes of cold acetone containing a 100-fold molar excess of unlabeled penicillin G. The resulting suspension was centrifuged at 11,000 X g for 20 min, and the pellet was dried under a stream of nitrogen.Denaturation, Reduction, Carboxymethylation, and Trypsin or Pronase Digestion. The precipitated carboxypeptidase, labeled with penicillin G (160 nmol, 1.1 X 107 dpm) was dissolved in 2 ml of 6 M guanidine-HCI containing 2 mM EDTA, and 0.2 M Tris-HCI (pH 8.2). After 6 mg of dithiothreitol had been added, the solution was incubated under nitrogen at 370 for 1 hr. Then 20 mg of iodoacetate was added and the solution was incubated under nitrogen and in the dark at room temperature for 1 hr. The solution was dialyzed against distilled water at 40 during which the carboxypeptidase precipitated. The resulting suspension was lyophilized. After lyophilization, the carboxymethylated carboxypeptidase was dissolved in 3 ml of 0.1 M NH4HCO3 and TPCK-trypsin was added to a final concentration of 10% (wt/wt) relative to the carboxypeptidase. The hydrolysis was carried out at 370 for 1 hr, and the reaction was stopped by freezing with dry ice-acetone followed by lyophilization. Pronase digestion was performed under the same conditions as trypsin digestion. If longer incubation times were used, much of the bound penicillin was released.

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The Source of Synovial Fluid Alkaline Phosphatase.

Cited by 9 publications

References 1 publication

Alkaline phosphatase activity associated to a calcium binding glycoprotein from calf scapula cartilage

Alkaline phosphatase activity associated to a calcium binding glycoprotein from calf scapula cartilage

An enzymatic method for determining magnesium

Isolation of the penicillin-binding peptide from D-alanine carboxypeptidase of Bacillus subtilis.

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