[99] Determination of nucleic acids in tissues by pentose analysis

Schneider, Walter

doi:10.1016/s0076-6879(57)03442-4

Cited by 1,751 publications

(648 citation statements)

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“…The concentration of DNA was determined spectrophotometrically at 600nm with diphenylamine reagent as described by Schneider (1957). The standard curve (50-800,ug of DNA) was prepared by using calf thymus DNA.…”

Section: Animalsmentioning

confidence: 99%

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Metabolic adaptations during lactogenesis. Lactose synthesis in rabbit mammary tissue during pregnancy and lactation

Mellenberger

Bauman

1974

Biochemical Journal

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1. Mammary tissue, obtained from rabbits at various stages of pregnancy and lactation, was used for tissue-slice incubations to measure the rate of lactose biosynthesis and for relevant enzymic activity measurements. A biphasic adaptation in the rate of lactose synthesis and in the RNA concentration was noted during lactogenesis. 2. The first increase in the rate oflactose biosynthesis occurred between days 15 and 24 ofpregnancy. A second substantial increase was noted immediately postpartum. The overall rate of lactose biosynthesis increased 12-fold from day 24 of pregnancy to day 15 of lactation postpartum, and then decreased from 15 to 22 days post partum. 3. The RNA concentration/g wet wt. of tissue and the ratio of RNA/DNA closely represented the biphasic ability of the mammary-tissue slice to synthesize lactose. 4. Increases in the activities of UDP-glucose 4-epimerase (EC 5.1.3.2) and lactose synthase (EC 2.4

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Section: Animalsmentioning

confidence: 99%

“…The standard curve (50-800,ug of DNA) was prepared by using calf thymus DNA. The concentration of RNA was determined with orcinol reagent (Schneider, 1957 (1957).…”

Section: Animalsmentioning

confidence: 99%

Metabolic adaptations during lactogenesis. Lactose synthesis in rabbit mammary tissue during pregnancy and lactation

Mellenberger

Bauman

1974

Biochemical Journal

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“…DNA was used as marker for nuclear materials and RNA for roughsurfaced endoplasmic reticulum (RER). The samples were extracted for nucleic acids as described by Schneider (1957), except for the use of perchloric acid instead of TCA, and RNA was determined by the orcinol reaction, while DNA was measured by the diphenylamine method of Giles and Myers (1965). Yeast RNA and calf thymus DNA were used as standards.…”

Section: Methodsmentioning

confidence: 99%

Plasma membrane associated enzymes of mammary tumours as the biochemical indicators of metastasizing capacity. Analyses of enriched plasma membrane preparations

Chatterjee¹,

Kim²,

Bielat³

1976

Br J Cancer

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Summary.-Plasma membranes from 6 spontaneously metastasizing and 4 nonmetastasizing rat mammary carcinomata were isolated by discontinuous sucrose density gradient centrifugation of microsomal pellets. The starting microsomal fraction contained 40-50%o plasma membranes as determined by the levels of 5'-nucleotidase activity, with a negligible amount of nuclear (1%), mitochondrial (5%o) and lysomal (7%o) contamination. Five distinct fractions (F1-F5) were banded at densities 1 09, 1 13, 1 15, 1-17 and 1-21 at 25°C, in addition to a pellet (F6) obtained by centrifuging at 76,000 g for 17 h. The fractions Fl through F5, all contained various concentrations of membranous structures, while the pellet (F6) contained only amorphous materials as evidenced by electron microscopy. The F3 fraction at the gradient 1 15 had the highest specific as well as total activity of the plasma membrane marker enzyme, with aggregates of the least contaminated plasma membranes in vesicular forms. This fraction also had the lowest specific activity for glucose-6-phosphatase (smooth ER marker) and for ,B-D-glucuronidase (lysomal marker), and therefore was considered to be the " cleanest " plasma membrane fraction. When the activity of 4 additional plasma membrane marker enzymes, i.e., alkaline phosphatase, phosphodiesterase I, nucleotide pyrophosphatase and alkaline ribonuclease was determined in the same F3 fraction, their levels were significantly lower in every metastasizing tumour than in the non-metastasizing ones, with the enzyme activity decreasing in direct proportion to the metastasizing capacity. On the other hand, the marker enzymes were high in all non-metastasizing tumours, with the activity seemingly increasing with the immunogenicity of tumour cells. There was no significant difference between the 2 groups of mammary tumours in the levels of sialic acid, hexosamine, phospholipid or cholesterol in the plasma membranes. Thus, the level of plasma membrane marker enzymes is considered an accurate indicator for metastasizing capacity in the rat mammary tumour system. IN EARLIER studies (Kim and Pickren, 1974;Kim et al., 1975), it was demonstrated that the level of various plasma membrane marker enzymes correlated roughly with the amount of membrane bound glycoproteins or glycocalyx and the immunogenicity of rat mammary tumour cells, and, inversely, with their antigen shedding property and metastasizing capacity. Since subcellular contaminants tend to interfere with an accurate quantitation of tumour cell plasma membranes, we have directed our efforts towards developing a method whereby the plasma membranes of rat mammary tumour cells, free of contaminations, can be prepared reproducibly. This paper describes an improved method for isolation of plasma membranes and also confirms and extends our earlier

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“…Ribose was determined by the orcinol method (Schneider, 1957). Ribitol was extracted from fresh mycelia by the method of Maruo et al (1965) and was separated and isolated by paper chromatography with butan-1-olpyridine-water (6: 4: 3, by vol.)…”

Section: Methodsmentioning

confidence: 99%

Ribitol and flavinogenesis in Eremothecium ashbyii

Mehta

Mattoo

Modi

1972

Biochemical Journal

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1. Supplementation of cultures of Erenwthecium ashbyii with ribitol leads to a twofold increase in riboflavin formation compared with unsupplemented cultures or those supplemented with ribose or ribulose phosphate. Addition of unlabelled ribitol decreases the incorporation of [1-14C]ribose into riboflavin, indicating that free ribitol is preferred to ribose for incorporation into riboflavin. 2. The enzymes ribitol kinase, D-ribose reductase, D-ribose 5'-phosphatase and GMP nucleosidase were demonstrated in the cell-free extracts. Ribitol induces the formation of ribitol kinase. The enzyme is activated in vitro by the flavinogenic purines, guanine and xanthine. D-Ribose reductase shows a specific requirement for NADPH and forms free ribitol from ribose. 3. The activities of ribitol kinase, ribose 5'-phosphatase and GMP nucleosidase reach their maximal values before riboflavin formation reaches a maximum. 4. [U-_4C]GMP is taken up intact by the culture ofE. ashbyii and is incorporated into riboflavin as well as into a blue fluorescent compound. The radioactivity from this compound is incorporated into riboflavin by the cell-free extract of E. ashbyii.Micro-organisms have been used extensively to study the biosynthesis of riboflavin, since some of them produce this vitamin in large excess. Although considerable information is available on the biosynthesis of riboflavin, the mode of insertion of the ribitol moiety into the vitamin molecule still remains obscure (Bacher & Lingens, 1970;Baugh & Krumdieck, 1969;Goodwin, 1963; Goodwin & Jones, 1956;Harvey & Plaut, 1966;Maley & Plaut, 1959;McNutt, 1954;Plaut, 1961). The present investigation was undertaken to study the effect of availability of ribitol and guanine on enzymic constitution and flavinogenesis in Eremothecium ashbyii. Materials and Methods OrganismA flavinogenic strain of Eremothecium ashbyii (no. 0944), obtained from the Type Culture Collection, Tokyo, Japan, was maintained on Sabourauds' agar (Difco Laboratories, Detroit, Mich., U.S.A.) slant cultures. The yeast was cultivated in the liquid fermentation medium as described by Goodwin & Pendlington (1954), supplemented with 0.5% vitamin-free Casamino acids (Difco Laboratories). The inoculated flasks were incubated at 28 ±20C in a dark incubator; unless otherwise specified, the cultures were incubated for 4 days. The surface cultures grown in the above liquid medium ('normal medium') are termed 'normal cultures'; when ribose (5 or lOmg/ 50ml) or ribitol (5 or lOmg/50ml) or guanine (3 mg/50ml) was included in this medium, the culVol. 130 tures are referred to as 'ribose-supplemented', 'ribitol-supplemented' or 'guanine-supplemented' respectively. Radioactivity experimentsTo study the fate of [1-14C]ribose during growth and flavinogenesis, the organisms were grown for 48h in the non-radioactive normal medium. Cells were collected by centrifugation and resuspended in the fermented broth to give a concentration ofapprox. 0.2mg dry wt./ml of suspension. Samples (5ml) from this suspension were distributed in lOOm...

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[99] Determination of nucleic acids in tissues by pentose analysis

Cited by 1,751 publications

References 13 publications

Metabolic adaptations during lactogenesis. Lactose synthesis in rabbit mammary tissue during pregnancy and lactation

Metabolic adaptations during lactogenesis. Lactose synthesis in rabbit mammary tissue during pregnancy and lactation

Plasma membrane associated enzymes of mammary tumours as the biochemical indicators of metastasizing capacity. Analyses of enriched plasma membrane preparations

Ribitol and flavinogenesis in Eremothecium ashbyii

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