The chemical fractionation of lymphatic organs

A method for chemically fractionating lymphatic organs has been described. The method has been shown to be applicable to bovine palatine tonsils, sheep palatine tonsils, and bovine thymus. Approximately 50 per cent of the dry weight of tonsils and about 30 per cent of thymus has been found to be soluble in the 0.15 M NaCl extract. Four components have been isolated which together account for 65 per cent by weight of the material in the extracts. Four other components have been identified and partially defined by means of electrophoretic mobility, solubility, or some other chemical or physical property.     

Calf thymus composition: a comparison of differential centrifugation and chemical fractionation procedures

The extraction behavior of thymus and the composition of fractions prepared from this organ has been studied. Sequential extraction methods using 0.15 M NaCl followed by water gave information with respect to the weight fraction of cytoplasmic and nuclear constituents. Lipide, nucleic acid, and electrophoretic analysis of the extracts provided additional information. A less complex electrophoretic pattern was obtained from subsequent extracts in the sequence. Sucrose and saline dispersates obtained from tissue fragmented with either the Potter-Elvehjem homogenizer or in a Waring blendor were fractionated, using standard differential sedimentation methods. The fractions obtained by means of four different dispersion procedures were compared in terms of yield, chemical analysis, and electrophoretic composition. The quantity of material in thymus having the sedimentation characteristics of liver mitochondrial and microsomal fractions was remarkably small. Both the suspension medium employed and the method used to bring about a disruption of the cells in the tissue affected the yield of "particulate" material. The components present in the later extracts in the sequence, E(4) to E(7), in the case of sequential extraction study resembled with respect to chemical composition and electrophoretic characteristics, the microsome fraction prepared by differential sedimentation methods. About 76 per cent of the PNA in the tissue appeared to be in the cytoplasm. The remaining 24 per cent PNA was found in the nucleus and accounted for 1.7 per cent of nucleus on a dry weight basis. From 75 to 88 per cent of cytoplasmic PNA was extracted from the tissue and 76 to 94 per cent of the PNA in the extract was found in the final supernatant solutions, depending upon the dispersion methods and suspension medium used in the extraction procedure. The composition of the final supernatant fractions using differential sedimentation methods were comparable in terms of electrophoretic properties, protein concentration, nucleic acid content, and fractionation behavior to saline extracts E(1) to E(3), of thymus used in earlier studies.     

STUDIES ON BIOLUMINESCENCE : II. THE PARTIAL PURIFICATION OF CYPRIDINA LUCIFERIN

Some solubility, oxidation, reduction, and compound-forming characteristics of extracts of Cypridina luciferin have been presented. A method of purification has been described which increased the amount of luciferin per unit of dry weight, as measured by the total light emitted, to about two thousand times that in the dry starting material. The best yields were from 50 to 65 per cent.     

Two proteins purified from lobster nerve extract; isolation and physicochemical characterization

1. A protein component, fraction B, of lobster nerve extracts has been isolated and purified by differential ultracentrifugation and precipitation with zinc acetate. 2. Physicochemical data obtained from this protein and from fraction C are summarized. 3. Fraction B is present in lobster nerve extracts in higher concentration (relative to fraction A) than in blood. 4. A second component, fraction C, of sedimentation constant S(20) (o) = 13.2 has been isolated from lobster nerve extracts.     

MICROSOMAL NUCLEOPROTEIN PARTICLES FROM PEA SEEDLINGS

Ultracentrifugal analysis of an extract of pea epicotyls, previously freed of debris and larger particles by centrifugation at 40,000 g for 10 minutes, has revealed the presence of a major component which possesses a sedimentation coefficient of 74 S. This component constitutes about 25 per cent of the TCA-precipitable material in the clarified epicotyl extract and is estimated to make up 1 to 2 per cent of the dry weight of the original tissue. In size, chemical composition, and morphology, the 74 S component resembles the nucleoproteins of the microsomes from animal tissues. The 74 S component of pea epicotyl extracts has been purified by repeated cycles of differential centrifugation to yield a preparation which is 80 per cent homogeneous in the analytical ultracentrifuge. It has been found to contain 30 to 37 per cent RNA as judged by a variety of analytical techniques. Approximately 55 per cent of the weight of the material is protein and a further 4.5 per cent phospholipide. Electron micrographs of air-dried specimens of the purified preparation show the 74 S constituent to be flattened spheres with an average height of 180 A and an average diameter of approximately 280 A. The molecular weight of the 74 S particles is computed from sedimentation, viscosity, and partial specific volume data to be 4.5 million ± 10 per cent in agreement with the value estimated from electron micrographs. The 74 S or microsomal component of pea epicotyls is rapidly aggregated in the presence of low concentrations of Mg ions or by somewhat higher concentrations of Ca or K salts. ATP on the contrary causes resolution of electrolyte-induced microsomal aggregates with simultaneous degradation of the particles to an ultracentrifugally inhomogeneous mixture of lower molecular weight materials.     

Male accessory gland substance of Melanoplus sanguinipes: An oviposition stimulant under the control of the corpus allatum

The accessory glands of male Melanoplus sanguinipes contain an oviposition stimulant. Injection of gland extracts from mature males induces oviposition in 75 per cent of capable virgin females within 24 hr. Injection of gland extracts from allatectomized males produces no stimulatory effect. Gland extracts from mature males contain two antigens which cannot be detected in gland extracts from allatectomized males. However, both antigens can be detected in gland extracts from allatectomized males 3 days after treatment with juvenile hormone.Anion-exchange chromatography of mature gland extracts yielded two fractions which, when injected into virgin females, induced oviposition in 71 per cent of capable insects within 24 hr. These two fractions are immunologically identical to the two antigens which are absent from gland extracts of allatectomized males. We suggest that synthesis of the oviposition stimulant in Melanoplus is controlled by the corpus allatum.Injection of brain extract also induces oviposition in 100 per cent of capable virgin females within 24 hr. A possible rôle for the brain in the oviposition process is discussed.     

Colorimetric analysis with N,N-dimethyl-p-phenylenediamine of the uptake of intravenously injected horseradish peroxidase by various tissues of the rat

1. A method is described for the colorimetric determination of peroxidase with N,N-dimethyl-p-phenylenediamine. The amount of red pigment formed by peroxidase is proportional to the concentration of enzyme and to the time of incubation during the first 40 to 90 seconds. The influence of the concentration of enzyme, N,N-dimethyl-p-phenylenediamine, H(2)O(2), the time of incubation, pH, the temperature, and the possible interference by oxidizing and reducing agents of tissues has been tested. 2. The method has been used to follow the uptake of intravenously injected horseradish peroxidase by 18 different tissues of the rat over a period of 30 hours. The highest concentration of the injected tracer enzyme was found in extracts of kidney, liver, bone marrow, thymus, and spleen. Considerable amounts were taken up by pancreas, prostate, epididymis, and small intestine. Lower concentrations were found in extracts of lung, stomach, heart, and skeletal muscle, aorta, skin, and connective tissue. No uptake was observed by brain and peripheral nerve tissue. 3. Tissue homogenates containing high concentrations of the injected peroxidase, in general also showed high or average activity of acid phosphatase. 4. Six hours after intravenous administration, the liver contained 27 per cent, the kidney 12 per cent, and the spleen, 1.4 per cent of the injected dose. 5. Approximately 20 per cent of the injected peroxidase was excreted in the urine during the first 6 hours, and the concentration of peroxidase in blood serum and urine fell exponentially during this time. After 6 hours, only low concentrations were excreted in the urine but low enzyme activity was still detectable after 30 hours. Approximately 6 per cent of the injected dose was excreted in the feces from 6 to 20 hours after administration. 6. After feeding through a stomach tube, low concentrations of peroxidase were found in blood serum and urine. Considerable variations in the extent of absorption from the gastrointestinal tract were observed in individual rats.     

The Occurrence of Translocated Antibiotics in Expressed, Plant Sap

The details of a convenient laboratory press are presented anda procedure by which plant sap is readily obtained is described.The pressing procedure has been shown to release 53–74per cent. of the total plant water as expressed sap. Preliminary observations on the uptake and translocation ofantibiotics by higher plants obtained by examining expressedsap are described. It is shown that the assay of expressed sapprovides a measure of the griseofulvin and chloramphenicol concentrationin leaves of plants grown in solutions of these antibioticsthat does not differ significantly from the value obtained bythe assay of water extracts. Extraction of leaves from plants grown in griseofulvin solutionswith organic solvents demonstrates a much greater antibioticcontent than is indicated by the assay of expressed sap. Toexplain this difference a ‘free’ antibiotic fraction,obtainable in expressed sap or water extracts, and a ‘bound’fraction recovered from the leaf only by organic solvent extractionare postulated.     

An evaluation of anti-multiplier factors in kidneys with obstructed ureters

Ureteral obstruction has been known to reduce the renal concentration gradients of chloride, sodium, urea, and osmolality. A quantitative analysis of the factors responsible for the decrease in the gradients has been performed. By applying the equation of conservation of matter to data obtained in this laboratory it is concluded that:a. at least, 62.5 per cent of the decrease in concentration gradients is due to drainage by circulatory vessels;b. at most, 25 per cent of the decrease is due to increase in water content of medulla; andc. at most, 12.5 per cent of the decrease is due to diffusion of solutes through the interstitium. Work done under the tenure of an Established Investigatorship of the American Heart Association.     

Free Amino Acids in the Leaves of Salvinia natans and Azolla filiculoides Grown in Light and Dark

Free amino acids were determined quantitatively by thin-layer chromatography and identified by various chromatographic methods in the leaves of two floating water ferns. Special interest was paid to the amount of γ-amino butyric acid. Water soluble nitrogen compounds increase in the leaves of both plants during the growth of three to four weeks in darkness. Compounds especially accumulating are ammonia and the amides asparagine and glutamine. The amount of most proper amino acids is reduced in the dark, except γ-amino butyric acid, which, in addition to amides, is the main amino compound increasing in darkness. 0.1 M HCl extracts the greatest amount of nitrogen compounds of low molecular weight from the dried leaves of the experimental plants. If we denote the amount of the dissolved amino compounds as 100 per cent, we can say that water extracts 96 per cent of the amino compounds, and that an 80 per cent ethanol solution extracts 79 per cent of these compounds.     

Colorimetric Analysis with N,N-Dimethyl-p-phenylenediamine of the Uptake of Intravenously Injected Horseradish Peroxidase by Various Tissues of the Rat

1. A method is described for the colorimetric determination of peroxidase with N,N-dimethyl-p-phenylenediamine. The amount of red pigment formed by peroxidase is proportional to the concentration of enzyme and to the time of incubation during the first 40 to 90 seconds. The influence of the concentration of enzyme, N,N-dimethyl-p-phenylenediamine, H₂O₂, the time of incubation, pH, the temperature, and the possible interference by oxidizing and reducing agents of tissues has been tested. 2. The method has been used to follow the uptake of intravenously injected horseradish peroxidase by 18 different tissues of the rat over a period of 30 hours. The highest concentration of the injected tracer enzyme was found in extracts of kidney, liver, bone marrow, thymus, and spleen. Considerable amounts were taken up by pancreas, prostate, epididymis, and small intestine. Lower concentrations were found in extracts of lung, stomach, heart, and skeletal muscle, aorta, skin, and connective tissue. No uptake was observed by brain and peripheral nerve tissue. 3. Tissue homogenates containing high concentrations of the injected peroxidase, in general also showed high or average activity of acid phosphatase. 4. Six hours after intravenous administration, the liver contained 27 per cent, the kidney 12 per cent, and the spleen, 1.4 per cent of the injected dose. 5. Approximately 20 per cent of the injected peroxidase was excreted in the urine during the first 6 hours, and the concentration of peroxidase in blood serum and urine fell exponentially during this time. After 6 hours, only low concentrations were excreted in the urine but low enzyme activity was still detectable after 30 hours. Approximately 6 per cent of the injected dose was excreted in the feces from 6 to 20 hours after administration. 6. After feeding through a stomach tube, low concentrations of peroxidase were found in blood serum and urine. Considerable variations in the extent of absorption from the gastrointestinal tract were observed in individual rats.     

REACTIONS TO INSECT ALLERGENS—The Incidence of Response to Testing Among Allergic and Nonallergic Persons

Positive reactions to skin tests with extracts of various insects were found to be significantly higher in 200 patients with allergic disease than in 150 nonallergic controls.Forty-seven per cent of the allergic group had positive reaction to bee, 46 per cent to house fly, 39.5 per cent to moth, 29.5 to ant and 28.5 to aphid. Ant and house fly extract caused the greatest number of severe reactions (4+), with moth second and bee third.Because extracts of nonstinging insects caused reaction almost as often and as strong as stinging insects, it may be inferred that sensitization is due to inhaled insect particles. There appeared to be no necessary connection between a history of having been stung and a strong skin test reaction to bee protein.Testing for insect sensitivity appears a good way to get diagnostic information that can enhance the effectiveness of treatment.     

Aerodynamic characteristics of the human right lung bronchial tree]

A new method of studying the aerodynamical characteristics of normal right bronchial tree in 17 natural preparations is described. The authors have found that 24 to 44 per cent of the total airflow entering the right main bronchus pass through the upper lobe bronchus, 9-26 per cent through the middle lobe bronchus and 38-56 per cent through the lower lobe bronchus. The data obtained show good agreement with the volume of the corresponding lobe expressed as percentage to the total right lung volume. Bronchial tree preparations differ greatly in their carrying capacities due to their corresponding aerodynamical resistances (+50% +25%). The relationship between resistance coefficient of the bronchial tree preparations and their geometrical parameters has been found.     

Role of lysyl-tRNA in the regulation of lysine biosynthesis in Escherichia coli K12.

A mutant of lysyl-tRNA synthetase has been isolated in Escherichia coli K12. With this strain the Kmapp for lysine is 25 fold higher than with the parental strain. The percentage of charged tRNAlys in vivo is only 7 per cent (as against 65 per cent with HFR H). Under these conditions no derepression of synthesis is observed for three lysine biosynthetic enzymes (AK III, ASA-dehydrogenase, DAP-decarboxylase) ; a partial derepression is obtained in the case of the dhdp-reductase. Thus lysyl-tRNA does not act as the only corepressor molecule in the lysine regulon.     

CESAREAN SECTION EXPERIENCE AT THE UNIVERSITY OF CALIFORNIA HOSPITAL

A review of the 746 cesarean sections performed at the University of California Hospital from 1907 through 1948 shows an overall cesarean section incidence of 3.91 per cent. The maternal mortality rate associated with these cesarean sections has been 1.61 per cent. There has been a steady decline in mortality over the 40-year period and there have been no deaths from cesarean section in the last ten years. The over-all incidence of morbidity associated with cesarean section has been 40.5 per cent and again there has been a significant improvement in recent years. The fetal mortality associated with cesarean section has been 6.1 per cent. In view of recent experience, standards based on figures collected 20 years ago are no longer tenable.     

THE COMBINATION OF GELATIN WITH HYDROCHLORIC ACID

The amount of HCl combined with a given weight of gelatin has been determined by hydrogen electrode measurements in 1 per cent, 2.5 per cent, and 5 per cent solutions of gelatin in HCl of various concentrations, by correcting for the amount of HCl necessary to give the same pH to an equal volume of water without protein. The curve so obtained indicates that the amount of HCl combined with 1 gm. of gelatin is constant between pH 1 and 2, being about 0.00092 moles.     

Studies on the Mechanism of Mold Protease Production

Inactive protease in the cell free extracts obtained from growing cells of Aspergillus sojae KS was collected in a supernatant of ultracentrifugation at 14×l0⁴ g, and in fractions obtained by acetone of 35~50 per cent and by ammonium sulfate of 0.5~0.6 saturation.

The inactive protease has the same resistance against pH or heat treatments as active protease has. The activation of the inactive protease was maximal between pH 5~6, and was accelerated by several kinds of protease, and was not affected by thioglycollate and KCN.     

Preparation,purification, and properties of E. coli virus T2

1. A method for the preparation of 8 to 10 liter quantities of T₂ virus lysates, titering 2 to 5 x 10¹¹ infectious units per ml. has been described. 2. Procedures have been developed for the concentration and purification of virus to a high specific infectivity. No fractionation procedure of the several used succeeded in further raising the specific infectivity of these purified preparations. 3. Some of the general properties of the better preparations have been determined. They exhibited titers of 2 x 10¹⁵ infective units per gm. of material or 1.2 x 10¹⁶ per gm. of nitrogen. 4. A study of the distribution of nitrogen among the various fractions of the virus showed that about 6 per cent of the total nitrogen is soluble in 4 per cent trichloracetic acid; that the protein nitrogen is about 40 per cent of the total and the nucleic acid nitrogen is 53 per cent. At least 96 per cent of the total phosphorus is in the nucleic acid fraction. Less than 0.5 per cent quantities of lipid and PNA were found.     

Presence and isolation of 2 lipoproteins immunologically related to apolipoprotein A I in human serum

Very high density lipoproteins d : 1.23--1.25 g/ml (VHDL2) have been isolated from human serum by preparative ultracentrifugation. They contain 80 per cent proteins and 20 per cent lipids. Lipids are mainly phospholipids (80 per cent). The proportion of lysolecithin (50 per cent) is higher than that of lecithin (40 per cent). The quantity of cholesterol is low, the free cholesterol: total cholesterol ratio is 0.35. VHDL2 consisted principally in lipoprotein D and two lipoproteins immunologically apparented to apolipoprotein A I, called LP A I1 and LP A I2. The LP A I1 has a molecular weight slightly higher and a hydrated density lower than that of LP AI2. Our experiments suggest that LP A I1 exists in the serum before ultracentrifugation while LP A I 2 comes from HDL degradation during ultracentrifugation. The immunological heterogeneity of apo A I forming different protein-lipid complexes is discussed.     

Effects of local hyperthermia on the motor function of the rat sciatic nerve

The effect of local heat treatment of the sciatic nerve was assessed using the toe-spreading test, which mainly assesses the motor function of the sciatic nerve. A 5 mm long segment of the nerve was heated at temperatures from 42.0 to 45.0 degrees C in vivo using a brass thermode. Hyperthermia led to a decrease in spreading of the toes. Recovery from functional loss took place in all cases, and this recovery was completed in 4 weeks. A 50 per cent functional loss in 50 per cent of the treated animals was observed after 58, 32 and 12 min of heating at 43.0, 44.0 and 45.0 degrees C respectively.