Supernatant fraction and

Figure 1. Top Turbidity, measured at 350 nm, as a function of microtubule polymer mass concentration (expressed as mg/mL polymerized tubulin). Tubulin solutions of varying concentrations were polymerized until they reached stable plateau values in a Cary 118C spectrophotometer. Each sample was then transferred to an ultracentrifuge tube, and microtubules were pelleted, separated from the unpolymerized tubulin in the supernatant fraction, and then resuspended for protein concentration determination. The corresponding turbidity and polymer mass concentrations are plotted here. Bottom Time-course of tubulin polymerization assayed by turbidity.Repro-duced from MacNeal and Purich with permission from the American Society for Biochemistry and Molecular Biology.

In an effort to establish the nature of the acetogenic substrates, raw sludge was separated into solids and supernatant fractions and were tested individually as substrates in the digesting sludge. Most of the substrates responsible for the rapid formation of acetate were found in the water-soluble solids fraction (Figure 3), and no acetogenic activity was observed in the supernatant fractions. 

Spin Trapping and Measurement of Oy and HO Generated in Aqueous Buffer Solution of Supernatant Fractions and Total Suspension of PM... 

Seven renal glycoside hydrolases have been measured in four sub-cellular fractions prepared from highly inbred aglycosuric ( F-line) and glycosuric (XA-line) Chinese hamsters (Cricetulus griseus) a- and /3-D-Galactosidases were highest in the nuclear and supernatant fractions and both showed reduced activities in the XA animals. 

Remove the supernatant fraction and transfer to a new 1.5-mL tube. The supernatant fraction is used for protein capture by... 

The enzymatic hydrolysis of retinyl esters in liver has been examined in retinol-depleted, retinoic acid-supplemented rats (Harrison et al., 1979) as well as in normal rats (Prystowsky et al., 1981). In both studies, the enzyme retinyl palmitate hydrolase showed an unusual subcellular distribution, being found in the nuclear, mitochondrial, and supernatant fractions and displayed a large variability in activity from rat to rat. For additional information regarding the esterification and hydrolysis of retinol the reader is again referred to Chapters 7 and 10. 

The difficulty of determining the role of intermediates, free or enz5une-bound, in the biosynthesis of the penicillins and cephalosporin C has depended partly on the rapid turnover and very low concentrations of such substances in the mycelial cell and partly on the existence of permeability barriers which complicate the interpretations of results obtained when possible intermediates are added to the extracellular fluid. Some of these difficulties would be removed by a cell-free system in which all or part of the bios5mthetic pathways could be followed. Mycelium of the Cephalosporium sp. which has been disrupted by ultrasonic treatment in buffered saline is able to produce penicillin N and cephalosporin C at a rapid rate after a period of lag (Demain, 1963 Abraham et al, 1964). Centrifugation of these preparations yield a supernatant fraction and a sediment. The supernatant alone produces no antibiotics and the sediment alone relatively little. Thus it appears that the enzyme systems necessary for the production of penicillin N and cephalosporin C are present in the sediment and precursors in the supernatant. Further studies are necessary to determine how far, if at all, the permeability barriers of the normal cell are permanently broken down in the sediment fraction. 

Partially purified ODC preparations have also been obtained from tomato ovaries and cultured tobacco cells (Heimer and Mizrahi, 1982), tobacco callus (Tiburdo et al., 1985a), and Helianthus tuberosus tubers (D Orazi and Bagni, 1987). These ODC preparations were present in the respective supernatant fractions and probably represented cytosolic enzymes. On toe other hand, there appears to be a pelletable ODC in tobacco and tomato (Slocum et al., 1984) as well as oats (Young, 1984), probably representing particulate enzymes. 

Zinc. The 2—3 g of zinc in the human body are widely distributed in every tissue and tissue duid (90—92). About 90 wt % is in muscle and bone unusually high concentrations are in the choroid of the eye and in the prostate gland (93). Almost all of the zinc in the blood is associated with carbonic anhydrase in the erythrocytes (94). Zinc is concentrated in nucleic acids (90), and found in the nuclear, mitochondrial, and supernatant fractions of all cells. 

Plasma Add cold acetonitrile, centrifuge, separate supernatant, fractionate by reverse-phase HPLC and TLC GC/MS and PNMR No data No data Weiss etal. 1994... 

The aliquot containing 3% input, the entire eluted fraction, and the aliquot of 3% flow-through supernatant is mixed with 4 x sample loading buffer (1 M Tris-HCl [pH 6.8], 40% glycerol, 8% SDS, 0.06%... 

Indicine IV-oxide (169) (Scheme 36) is a clinically important pyrrolizidine alkaloid being used in the treatment of neoplasms. The compound is an attractive drug candidate because it does not have the acute toxicity observed in other pyrrolizidine alkaloids. Indicine IV-oxide apparently demonstrates increased biological activity and toxicity after reduction to the tertiary amine. Duffel and Gillespie (90) demonstrated that horseradish peroxidase catalyzes the reduction of indicine IV-oxide to indicine in an anaerobic reaction requiring a reduced pyridine nucleotide (either NADH or NADPH) and a flavin coenzyme (FMN or FAD). Rat liver microsomes and the 100,000 x g supernatant fraction also catalyze the reduction of the IV-oxide, and cofactor requirements and inhibition characteristics with these enzyme systems are similar to those exhibited by horseradish peroxidase. Sodium azide inhibited the TV-oxide reduction reaction, while aminotriazole did not. With rat liver microsomes, IV-octylamine decreased... 

Figure 4.7. Histamine release from mast cells in response to various dilutions of HRA generated from bovine serum albumin (BSA) by medium derived from stimulated rat neutrophils [156]. Neutrophils ((50-100) x K lml) were stimulated with FMLP (10 5 M), the medium removed and incubated with BSA (10 mg I ml) at pH 4.5 for 18 h at 37° C. It was then boiled and centrifuged (11,000 x g for 30 s), the supernatant fraction was removed, its pH was adjusted to 7.2 and it was added, at various dilutions, to suspensions of mast cells. Histamine release was then measured after 10 min. As the amount of generated HRA was increased histamine release increased to a maximum at 57 4%. Mean + S.E.M., n = 5. Inset Time-course of generation of HRA as assayed by histamine release from isolated mast cells. HRA was generated as before using 50 x 106 neutrophils/ml. Aliquots were removed at the indicated times and assayed (at 50% dilution) for HRA. Note that there is a significant generation of HRA by 2 h. Mean S.E.M., n = 3.

To acquire information on the intrinsic metabolic activity of aquatic organisms, liver of carp (Cyprinus carpio Linnaeus), rainbow trout (Salmo gairdneri) and freshwater snail (Cipango-paludina japonica Martens) was dissected out, homogenized in 0.1M phosphate buffer, pH 7.5, and centrifuged at 105,000 g for 60 min to obtain the microsome-equivalent (described as the microsomal fraction hereafter) fraction. The protein content of microsomal and submicrosomal (supernatant fractions by Lowry s method, microsomal P-450 content ( ), activity of aniline hydroxylase (4) and aminopyrine N-demethylase (5) were determined. 

FRACTIONATION AND ANALYSIS OF MS5 SUPERNATANT IDENTIFIES CSF-1 AS A MAJOR GROWTH FACTOR COMPONENT FOR MYL-D7 STEM CELLS... 

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