Luigi Messineo

A sensitive spectrophotometric method for the determination of free or bound tryptophan

Abstract 1. 1. A spectrophotometric method for the determination of tryptophan at 518 nm. is presented. 2. 2. The method is simple, fast (1 hour), sensitive (1.20 μg. tryptophan), and is not interferred with by compounds that form non-specific colour products in sulphuric acid. 3. 3. The recovery of free and bound tryptophan in proteins is within experimental error.

Sensitive spectrophotometric determination of fructose, sucrose, and inulin without interference from aldohexoses, aldopentoses, and ketopentoses

Abstract 1. 1. Two spectrophotometric methods for the determination of fructose, sucrose, fructose phosphates, and inulin as a function of fructose content are presented. 2. 2. Cysteine hydrochloride in 68–70 per cent (by volume) sulphuric acid is the reagent for the formation of a green chromophore (λmax, = 415 nm.); tryptophan, indole, and some of their derivatives complex with this chromophore to form another pink chromophore (λmax, = 518 nm.). 3. 3. The cysteine hydrochloride-tryptophan method is twice as sensitive as cysteine hydrochloride alone. One to 20 ng. of fructose can be determined. 4. 4. Both methods are better than previous methods, because they are specific, quick, and the chromophores are stable.

Transition metals in calf thymus deoxyribonucleoprotein

Fe, Ni, Cu and Zn were found by energy-dispersive X-ray fluorescence in calf thymus deoxyribonucleo-protein. The X-ray analyses indicated the absence of Cr, Mn and Co.

Age related changes in total DNA and RNA and incorporation of uridine and thymidine in rat brain

1. Total brain DNA and total brain RNA and the incorporation of thymidine[14C] and uridine[3H] were measured in young and aged rats. 2. From 20 days to the time of sexual maturation, both DNA and RNA levels increase. Total RNA exceeds total DNA at all ages. Comparatively, the ratio of total DNA/RNA is higher in young than in aged animals. 3. The incorporation of thymidine[14C]/g of DNA and of uridine[3H]/g of RNA decreases with age. This decrease is rapid in young animals. After 350 days of age, the incorporation becomes very low. The significance of data is discussed.

Temporal changes in histone acetylation.

The incorporation of tritiated acetate was studied in developing and aging rats. Thymus, liver and serum were collected 30 minutes after injection of acetate. The trichloroacetic acid precipitable histone fraction was then extracted from liver and thymus and its radioactivity determined. Serum and cytoplasmic fractions were also counted. Serum activity declined with age. Thymus histone and cytoplasmic fractions showed a cyclic pattern. Acetylation of liver histones showed a straight line decline to a relatively constant level. The decline in acetylation of liver histones is postulated to be due to repression of acetylation processes and is thought to parallel the change of hepatocytes from a diploid to a polyploid state.

Age-related changes in total dna and rna and incorporation of uridine and thymidine in rat liver, kidney and spleen

Total content of DNA and RNA in liver, kidney and spleen were measured in young and aged rats. At the same time the incorporation of [14C]thymidine, a DNA precursor, and [3H]uridine, an RNA precursor, were also determined. Changes in total organ DNA and RNA correlated with sexual maturation as did incorporation of precursors. Young animals have more DNA per organ relative to RNA, with kidney and spleen DNA showing a decrease between maturity and senescence. However, liver RNA increases with age, a change probably due to decreased catabolism of RNA since [3H]urine uptake decreases. Liver polyploid differentiation, and [14C]thymidine and [3H]uridine uptake, are correlated. In kidney, incorporation of [3H]uridine is inversely related to [14C]thymidine incorporation.

Characterization of sea-urchin sperm deoxyribonucleoproteins (DNP) extracted with glycine

Abstract 1. 1. Solubility, electrophoretic, ultracentrifugal, and electron microscopic evidence indicates that glycine-extracted DNP from sea-urchin sperm behaves as a mono-dispersed solution. 2. 2. If these mono-dispersed DNP solutions are exposed to salt solutions (o.o3–o.o7 M ) they give rise to three fractions. These fractions have a DNA to protein ratio of 30–70 per cent by weight respectively. These fractions are shown by electrophoresis. ultracentrifugation, and electron microscopy. 3. 3. In salt concentrations greater than 0–07 M more fractions are formed whose DNA to protein ratios are different, indicating a separation of proteins from DNA. 4. 4. The glycine extraction is compared with the salt extraction. There are strong indications that the usual washing of the nuclei and of the DNP with 0–1 M Nacl solutions damages the native DNP.

The effects of EDTA and contaminating metals on Fe, Cu and Zn in deoxyribonucleoprotein.

Abstract 1. 1. A substantial loss of native metals in DNP was observed when EDTA was present during cell homogenization or in the final extract. 2. 2. Similarly, the presence of trace amounts of Fe. Cu, and Zn resulted in metal contamination.

A test for ribose determination without interference from deoxyribose

Abstract 1. 1. The basic cysteine reaction for ribose, described by Dische in 1949, has been modified.In the present method the cysteine hydrochloride is added immediately after the addition of concentrated sulphuric acid. 2. 2. Thus the reaction time has been reduced from over 2 hours to 0–5 hour total time. 3. 3. The reaction is twice as sensitive as in the original Dische method, measuring solution of 10–60 μg.of ribose. 4. 4. The reaction has also been modified in the blank used: in the present method it does not contain cysteine hydrochloride but it contains the test solution. 5. 5. In the original Dische method DNA, deoxyribose, and many other compounds interfered.We have found that deoxyribose and DNA give a non-specific yellowish colour in the presence of sulphuric acid, the intensity of which is not affected by the presence of cysteine hydrochloride.Thus, the use of a blank containing the same test solution in sulphuric acid but without cysteine hydrochloride eliminates the interference due to deoxyribose, DNA, and other compounds such as proteins.

Ionic strength effects on nucleoproteins

Abstract 1. 1. Homogenization of calf (Bos bovinus) thymus cells in 0.15 M triethanolamine hydrochloride (TEA-HC1) prior to the extraction of deoxyribonucleoprotein (DNP) with 0.01 M glycine results in the recovery of a fractionated DNP when compared to extracts from cells initially exposed to lower ionic strength TEA-HC1 solutions. 2. 2. The DNP from cells homogenized in 0.15 M TEA-HC1 contains approximately 10% less protein than the DNP from cells exposed to 0.007 M TEA-HC1.