The Na+ gradient-dependent transport of D-glucose in renal brush border membranes.

The Na+-dependent transport of D-glucose was studied in brush border membrane vesicles isolated from the rabbit renal cortex. The presence of a Na+ gradient between the external incubation medium and the intravesicular medium induced a marked stimulation of D-glucose uptake. Accumulation of the sugar in the vesicles reached a maximum and then decreased, indicating efflux. The final level of uptake of the sugar in the presence of the Na+ gradient was identical with that attained in the absence of the gradient, suggesting that equilibrium was established. At the peak of the overshoot the uptake of D-glucose was more than 10-fold the equilibrium value. These results suggest that the imposition of a large extravesicular to intravesicular gradient of Na+ effects the transient movement of D-glucose into renal brush border membranes against its concentration gradient. The stimulation of D-glucose uptake into the membranes was specific for Na+. The rate of uptake was enhanced with increased concentration of Na+. Increasing Na+ in the external medium lowered the apparent Km for D-glucose. The Na+ gradient effect on D-glucose transport was dissected into a stimulatory effect when Na+ and sugar were on the same side of the membrane (cis stimulation) and an inhibitory effect when Na+ and sugar were on opposite sides of the membrane (trans inhibition). The uptake of D-glucose, at a given concentration of sugar, reflected the sum of the contributions from a Na+-dependent transport system and a Na+-independent system. The relative stimulation of D-glucose uptake by Na+ decreased as the sugar concentration increased. It is suggested, however, that at physiological concentrations of D-glucose the asymmetry of Na+ across the brush border membrane might fully account for uphill D-glucose transport. The physiological significance of the findings is enhanced additionally by observations that the Na+-dependent D-glucose transport system in the membranes in vitro possessed the sugar specificities and higg phlorizin sensitivity characteristic of more intact preparations. These results provide strong experimental evidence for the role of Na+ in transporting D-glucose across the renal proximal tubule luminal membrane.


Bovine kidney alkaline phosphatase. Purification, subunit structure, and metalloenzyme properties.

Kidney alkaline phosphatase was purified to homogeneity. It is a glycoprotein of about 172,000 molecular weight. Analyses of the subunit structure by sedimentation equilibrium in 6 M guanidine hydrochloride and by gel electrophoresis in sodium dodecyl sulfate indicate that the alkaline phosphatase is a dimer comprising two very similar or identical subunits of about 87,000 molecular weight. The native enzyme contains 4.5 +/- 0.2 g atoms of zinc per mol of protein. Reconstitution experiments from the apophosphatase show that binding of 4 Zn2+ per mol of dimer is essential for full activity. The kinetic data of Zn2+ binding to the apoprotein require at least a two-step mechanism, in which one of the steps corresponds to a conformational change within the enzyme. This paper also presents data concerning amino acid composition, sugar content, enzyme stability, absorbance index, and sedimentation velocity.


Changes in globin messenger RNA content during erythroid cell differentiation.

Previous studies have shown that mouse fetal erythroid precursor cells isolated by an immunological technique synthesize little or no globin and contain little, if any, globin mRNA, as assayed in a cell-free system (translatable mRNA). After culture for 10 hours in the presence of erythropoietin, there is a marked increase in globin synthesis and in translatable globin mRNA. The present studies were designed to measure directly the content of globin mRNA sequences during erythroid cell differentiation, by molecular hybridization with 3H-labeled DNA complementary to globin mRNA. The results indicate that few, if any, globin mRNA sequences are present in the total RNA of erythroid precursor cells. There is little or no pool of untranslated globin mRNA in these cells. After 10 hours of culture with erythropoietin, there is an increase in globin mRNA content, as ;easured by a change in the Cot1/2 values obtained by cDNA: mRNA hybridization with (Co) representing the concentration of RNA. Between 0 and 22 hours of culture, there is a 250-fold rise, and between 22 and 44 hours, a further 2-fold increase in globin mRNA content. During the 44 hours in culture, the number of cells in culture increases 2- to 3-fold. The number of globin mRNA molecules rises in erythroid precursor cells to an average value of 1800 molecules/cell during 22 hours of culture. In cultures without added erythropoietin, the absolute number of cells decreases, however, cells presumably induced to differentiate by exposure to erythropoietin in vivo continue to differentiate in vitro, accumulating globin mRNA and initiating globin synthesis.


Improved methods for purification and assay of eukaryotic messenger ribonucleic acids and ribosomes. Quantitative analysis of their interaction in a fractionated reticulocyte cell-free system.

The polyadenylic acid-containing messenger ribonucleic acids of eukaryotic cells are rapidly isolated and deproteinized in a simple and gentle one-step procedure. The polyribosome fraction, dissolved in 0.5 M NaCl/0.5 percent sodium dodecyl sulfate, is passed through an oligo(dT)-cellulose column which is then washed with the solvent until proteins and contaminating ribonucleic acids are fully removed. Deproteinized messenger ribonucleic acid is then eluted by lowering the ionic strength. This method gives highly purified and active messenger ribonucleic acids from all tissues tested. The yield is approximately 1.5 to 2 percent of the polyribosomal ribonucleic acid. Messenger ribonucleic acids are assayed in a rabbit reticulocyte-derived, messenger-dependent, cell-free protein-synthesizing system modified from Crystal et al. (Crystal, R. G., Nienhuis, A. W., Elson, N. A., and Anderson, W.F. (1972) J. Biol. Chem. 247, 5357-5368). This system synthesizes proteins at an almost linear rate for at least 2 hours. During this period, each globin messenger ribonucleic acid directs the synthesis of several globin molecules. Each active ribosome synthesizes a globin molecule every 6 to 7 min, but only a small fraction of the ribosomes or messengers are active at any instant. Translation occurs mainly on di- and monoribosomes although larger sized polysomes also occur. Several lines of evidence suggest that globin messenger ribonucleic acid requires "activation" before it can be utilized and that a messenger activation step of protein synthesis initiation is rate-limiting in this cell-free system.


Potentiation of hemoglobin messenger ribonucleic acid. A step in protein synthesis initiation involving interaction of messenger with 18 S ribosomal ribonucleic acid.

The polyadenylic acid-containing messenger ribonucleic acid from rabbit reticulocyte polyribosomes, isolated by a rapid and very gentle procedure (Krystosek, A., Cawthon, M. L., and Kabat, D. (1975) J. Biol. Chem. 250, 6077-6084), sediments in a sucrose gradient in three sharp peaks, at 9 S, 17 to 18 S, and 28 S. The alpha and beta globin messenger activity follows the absorbance profile in the sucrose gradients and has its major peak at 17 to 18 S. The larger messengers are more active than 9 S messenger by approximately 2-fold per mass unit of ribonucleic acid or by at least 8-fold per molecule. The major 17 to 18 S form of globin messenger was examined further and was shown to be a 1:1 complex of 9 S messenger and 18 S ribosomal ribonucleic acid. The effect of 18 S ribosomal ribonucleic acid on translation of purified 9 S globin messenger was analyzed in a messenger-dependent protein-synthesizing system (Krystosek, A., Cawthon, M. L., and Kabat, D. (1975) J. Biol. Chem. 250, 6077-6084). In the absence of exogenous ribosomal ribonucleic acid, 9 S messenger is inefficiently translated; a large excess of messenger is required to saturate the system; and globin is synthesized mainly on di- and monoribosomes. Exogenous liver or reticulocyte 18 S ribosomal ribonucleic acid potentiates 9 S messenger translation and renders it at least 10 times more efficient. The potentiation reaction can also be accomplished by increasing the concentration of ribosomes in the assay system. However, transfer or messenger ribonucleic acids cannot carry out this reaction. It is proposed that 9 S globin messenger ribonucleic acid is an inactive molecule which is normally potentiated by specific reversible base pairing with an accessible region of ribosomal ribonucleic acid contained in a 40 S ribosomal subunit. The potentiated messenger interacts with initiation factors and with other ribosomal subunits to synthesize protein. Potentiation is the first specific function in protein synthesis demonstrated for the ribosomal ribonucleic acid portion of ribosomes.


Molecular weight of the acetylcholine receptors of electric organs and the effect of Triton X-100.

Studies are reported on the influence of Triton X-100 on the molecular weight and functional properties of the acetylcholine receptor. Results are presented principally for receptors purified from Torpedo californica and Torpedo marmorata with a limited number of observations on the receptor from Electrophorus electricus. In equilibrium dialysis measurements Trito, X-100 greatly reduced acetylcholine binding to the high affinity sites of the receptor from T. californica, but had only a small effect on the sites of lower affinity. Sedimentation equilibrium experiments on receptor in the absence of added Triton X-100 revealed average apparent molecular weight values of 510,000 for receptor from T. californica and 665,000 for T. marmorata. Under those conditions 0.113 mg of residual Triton X-100 were found per mg of protein as determined by using 3H-labeled Triton X-100. The sedimentation data indicated the presence of more than one molecular species, involving a unit with an apparent molecular weight of 330,000 and higher aggregates. Upon addition of Triton X-100, the higher aggregates were reduced, and above 0.1 percent Triton X-100 the 330,000 unit was the principal component present for receptor from all three species examined. Various structural models are considered in the light of this value, the polypeptide size from Na dodecyl sulfate-gel electrophoresis, and the protomer size determined by the molecular weight of an acetylcholine binding site.


Retinoic acid-binding protein in rat tissue. Partial purification and comparison to rat tissue retinol-binding protein.

When the 100,000 X g supernatant fractions of several rat organs are incubated with all-trans-[3H]retinoic acid, a binding component for retinoic acid with a sedimentation coefficient of 2 S can be detected by sucrose gradient centrifugation. This tissue binding protein for retinoic acid is distinct from the tissue binding protein for retinol which has been previously described. The tissue retinoic acid-binding protein has been partially purified from rat testis and this partially purified protein would appear to have a molecular weight of 14,500 as determined by gel filtration and high binding specificity for all-trans-retinoic acid. Binding of [3H]retinoic acid is not diminished by a 200-fold molar excess of retinal, retinol, or oleic acid but is reduced by a 200-fold excess of unlabeled retinoic acid. Tissue retinoic acid-binding protein can be detected in extracts of brain, eye, ovary, testis, and uterus but is apparently absent in heart muscle, small intestine, kidney, liver, lung, gastrocnemious muscle, serum, and spleen. This distribution is different than that observed for the tissue retinol-binding protein. Tissue retinol-binding protein was also purified extensively from rat testis. The partially purified protein has an apparent molecular weight of 14,000 and high binding specificity for all-trans-[3H]retinol as only unlabeled all-trans-retinol but not retinal, retinoic acid, retinyl acetate, retinyl palmitate, or oleic acid could diminish binding of the 3H ligand under the conditions employed. The partially purified protein has a fluorescence excitation spectrum with lambda max at 350 nm. In contrast, the retinol-binding protein isolated from rat serum and described by others has a fluorescence excitation spectrum with lambda max at 334 nm and an apparent molecular weight of 19,000. When partially purified tissue retinol-binding protein is extracted with heptane, the heptane extract has a fluorescence excitation spectrum similar to that of all-trans-retinol.


Restoration of function in the knee and elbow with a hinge-distractor apparatus.

External hinged distraction devices using transosseous pins and adjustable attachments were applied to mobilize joint contractures; to reduce old dislocations; and to reduce, immobilize, and compress ununited para-articular fractures. Once these procedures were accomplished the devices were then used to maintain the joint surfaces separated at predetermined distances while kinematically normal joint motion was gradually restored, first passively and then actively. The results in thirty-one knees and twenty-eight elbows treated with these devices were encouraging after follow-ups of one to six years.


Preliminary experience with a method of quadricepsplasty in recurrent subluxation of the patella.

Nineteen knees were treated for recurrent subluxation or dislocation of the patella by lateral and distal transfer of the vastus medialis with or without release of the lateral retinaculum. Fifty-eight per cent has good or excellent and 42 per cent had poor results after a mean follow-up of twenty-nine months. The factors that predisposed to failure were genu valgum greater than 15 degrees and, to a lesser extent, palpable synovitis associated with symptomatic chondromalacia patellae.


Reconstruction of the anterior cruciate ligament by semitendinosus tenodesis.

Rupture of the anterior cruciate ligament constitutes a major disability in young athletes and those patients with persistent disabling instability who have failed to respond to conservative treatment. A new method of reconstruction by means of semitendinsous tenodesis was used in five patients with satisfactory end results after an average follow-up of 21.4 months. Limited clinical experience indicates that with this method less anteroposterior instability persists than with other methods.


Rotational control of unstable subcapital fractures. An experimental study.

An experimental evaluation of the stability of rotational fixation of unstable, subcapital femoral-neck fractures as provided by either the multiple pin and side-plated Deyerle device or the heavy duty Jewett triflanged nail showed no significant difference between the two devices. The determination of moment versus deformation behavior included study of two torsional axis orientations and superposition of simulated body-weight and muscular-force impaction loadings.


Preoperative skeletal traction in scoliosis.

To determine the role of preoperative traction in the treatment of spinal curves, ten patients with curves averaging 81 degrees by the Cobb measurement were studied. Each patient was treated in traction by means of a halo or skull tongs and femoral pins for two to three weeks prior to the surgical correction of the scoliosis with instrumentation. Weights were added to the traction apparatus, 1.8 kilograms a day, to a maximum of 18.1 kilograms. Pretraction supine roentgenograms were compared with supine roentgenograms made at three-day intervals throughout the traction period. Preoperative traction resulted in an average improvement of the curves of 34 degrees, or 41 per cent of the initial curve. Most of the correction was obtained within the first week of traction. Further correction during surgery with Harrington instrumentation yielded 13.5 degrees on the average (16 per cent) so that combined correction averaged 47 degrees, or 57 per cent.


Total joint replacement for ankylosed hips. Indications , technique, and preliminary results.

Sixteen patients with disabling pain in the back, knee, or hip had total hip-replacement arthroplasty. Good relief of pain, a limited but satisfactory range of motion (dependent on duration of ankylosis), and variable function resulted. Establishing active abduction was important in restoring function and achieving a negative Trendelenburg test. Careful preoperative assessment of abductor muscle strength, medialization of the acetabulum, use of long-neck prostheses with small and straight stems and good inherent stability, and muscle reconditioning were the keys to a good result.


Paralytic pelvic obliquity. Its prognosis and management and the development of a technique for full correction of the deformity.

Thirty-nine patients with post-poliomyelitic scoliosis and pelvic obliquity were treated and followed for up to four years. The dominant pathological mechanism causing the pelvic obliquity was trunk-muscle imbalance. The natural history of untreated pelvic obliquity is progression, leading ultimately to dislocation of the hip. The functional abilities of the patient are then severely restricted. Our aim in treatment was to obtain the maximum possible correction of pelvic tilt and so to prevent subluxation of the femoral head. Total correction was obtained in fifteen patients; no loss of correction was seen in twenty-four patients. The best results were achieved by preliminary traction, Dwyer instrumentation distally to the fifth lumbar vertebra, and extensive posterior fusion extendind distally to the sacrum.


Septic arthritis due to  pseudomonas in heroin addicts.

Ten cases of septic arthritis due to Pseudomonas occurred in users of heroin. Nine cases were monarticular, two each occurring in the sternoclavicular and the sacro-iliac joints. Review of the literature suggests that the incidence of Pseudomonas septic arthritis is increasing, especially among heroin addicts. The reported cases in adults suggest a predilection for the sternoclavicular joints (eight of forty-one) which is even more pronounced among addicts (seven of twenty-four). Pseudomonas infection associated with drug abuse should be suspected in cases of isolated sternoclavicular inflammation. The drugs used for treatment of septic arthritis due to Pseudomonas should include gentamicin and carbenicillin, in conjunction with adequate drainage by aspiration or surgery.


Clinical and experimental studies of the congenital constriction band syndrome, with an emphasis on its etiology.

The findings in fifty-nine patients with congenital constriction band syndrome and in experiments in which limb malformations resembling those of the human constriction band syndrome were successfully reproduced in rat fetuses by amniocentesis indicated that these malformations arise from excessive contraction of the uterine muscle during pregnancy, with resulting hemorrhages from the marginal blood sinuses of the digital rays. Such malformations in humans may arise during the fifth and sixth weeks counted from ovulation. It therefore was concluded that this syndrome is not hereditary but is produced by prenatal environmental factors.


Congenital dislocation of the hip associated with central core disease.

Central core disease is a congenital, non-progessive myopathy with proximal muscle weakness most prevalent about the pelvic girdle. Including out two patients  there have been seven recorded cases of congenital hip dislocation associated with central core disease in children. Initial and recurrent contractures about the hip and knee should suggest the possibility of central core disease. However, the definitive diagnosis can only be made by muscle biopsy.


Treatment of Legg-Calvé-Perthes disease. Prognostic value of Catterall's Classification.

Catterall's classification of Legg-Calvé-Perthes disease is of prognostic value. When indicated, early energetic treatment in the form of containment and achievement of a full range of motion significantly decreases the percentage of poor results.


Use of polymethylmethacrylate to enhance screw fixation in bone.

Pull-out testing of screws inserted into cement and bone under various conditions showed that the cement-screw complex was significantly stronger when the screw was placed in soft cement and the cement was allowed to polymerize without further manipulation. When screw fixation in osteoporotic bone was reinforced with cement, the bone was the weakest component in the system. Fixation under these conditions should be enhanced by increasing the area of contact between the cement and bone. By cooling the cement to prolong its working time, it could be injected with a syringe in such a way that maximum endosteal  and periosteal contact was provided.


Bone mineral: effects of oral contraceptives, pregnancy, and lactation.

Estimating bone mineral by the photon absorption (125I) method applied to the distal part of the radius, it was found that young women using oral contraceptives containing a daily dose of 100 micrograms of mestranol had higher concentrations of bone mineral than non-users. Women twenty to fifty-nine years old who had lactated were among the poorly mineralized, while those who had lactated but were now using oral contraceptives in various combinations were among the  highly mineralized.


Isolated avulsion of the tibial attachment of the posterior cruciate ligament of the knee.

Of fourteen isolated avulsion fracutres of the posterior tibial attachment of the posterior cruciate ligament, one was undisplaced and five were minimally displaced. These six were treated conservatively, but four of the five displaced avulsion fractures progressed to non-union and significant functional disability. Eight displaced avulsion injuries were treated by open reduction. In four knees the fragment was sutured back to its origin on the posterior superior aspect of the tibia, and three of the these united. In two knees the fragment was large enough to fix with a screw and union ensued. There was excellent functional capacity in five of the six that united. The other two displaced fractures, treated three and seven months after injury, had open reduction and the fragments were sutured to the tibia in an advanced position. Union was obtained in both with good functional capacity.


Plantar fasciitis. The painful heel syndrome.

Of 116 patients with pain in the plantar portion of the heel, nineteen proved on follow-up to have systemic disease as the etiology. Of these treated with phenylbutazone, 71 per cent showed good results and a similar percentage benefited equally from injections of cortisone derivatives. Only two patients required surgical procedures, and these were successful in both.


Ulnar variance in Kienböck's disease.

The distal extent of the radius and ulna (ulnar variance) was compared on roentgenograms of normal wrists in randomly selected black and white patients and of fifteen affected wrists in patients with Kienbock's disease. The results establish a statistically significant association between negative ulnar variance and Kienbock's disease. Blacks have more positive ulnar variance and the disease is less likely to develop in them than in whites.


Fracture of the medial condyle of the humerus.

Nine patients with ten fractures of the medial condyle of the humerus were reviewed after an average follow-up of 5.2 years. These avulsion fractures are unstable and there is a definite risk of nonunion. When the condylar fragment was rotated, open reduction and suture with catgut gave bad results. Rigid internal fixation is recommended.


Nailing versus prosthesis for femoral-neck fractures. A critical review of long-term results in two hundred and thirty-nine consecutive private patients.

In a follow-up study of 239 consecutive patients with fractures of the femoral neck treated by six surgeons in one hospital by Pugh nailing, Austin Moore prosthesis, or Knowles pinning, 96 per cent could be followed for a minimum of two years or until a definitve result had been reached. The incidence of unsatisfactory results was 39.3 per cent after Pugh nailing, 18.2 per cent after prosthetic replacement, and 14.2 per cent after Knowles pinning (mostly undisplaced or impacted fractures). Thirty-five unsatisfactory results following Pugh nailing were due to: aseptic necrosis which was definite in eighteen cases and probable in two; early mechanical failure in twelve cases; infection in one; and late non-union in two. Eight poor results followed prosthetic replacement and were due to erosion of the acetabulum in six and dislocation in two. The four poor results that followed Knowles pinning were due to aseptic necrosis in three and to pain, probably the results of aseptic necrosis, in another. From this study it was concluded that aseptic necrosis was more influenced by the original displacement than by the accuracy of reduction or fixation, and that in displaced femoral-neck fractures in the elderly, prosthetic replacement gave a more reliable result.


Mechanicla properties and mineral content of avascular and revascularizing cortical bone.

The mechanical properties of avascular and revascularizing bone and their mineral content were studied in thirty adult dogs. The strength and mineral content of avascular bone decreased in the early phase of avascular necrosis, and with revascularization the bone-mineral content increased but the mechanical strength decreased further. In the normal bone the mechanical strength increased with an increase in mineral content.


Microfractures in the head of the femur.

Microfractures of the trabeculae were studied in the femoral heads of three different population groups. The femoral heads were sliced and then examined with a dissecting microscope. Microfractures were found in large numbers in specimens from British patients who had suffered femoral-neck fracture, and in smaller numbers in femoral heads -aken routine autopsy in India and in China (Hong Kong). There were many more microfractures in the thinner, lightly stressed trabeculae than in the thicker  weight-bearing trabeculae in the fractured femoral-neck specimens. The significance of these findings is discussed in the light of current concepts regarding the pathogenesis and significance of trabecular microfractures.


A simplified technique for treating the complex dislocation of the index metacarpophalangeal joint.

The volar approach to open reduction of the complex dislocation of the index metacarpophalangeal joint as described by Kaplan proved to have certain disadvantages. Digital nerves are easily damaged during exposure and there is a limited view of the entrapped fibrocartilaginous volar plate dorsal to the metarcarpal head. A direct dorsal longitudinal incision through the skin and extensor tendon gives full exposure. The volar plate attached to the proximal phalanx and trapped over the dorsal aspect of the metacarpal head is in full view. The volar plate is split longitudinally and the dislocation reduces spontaneously as the flexor tendons and lumbrical muscle slip by the metacarpal head. The advantages of this approach as compared with the volar approach are: (1) there is full exposure of the fibrocartilaginous volar plate, the main structure blocking reduction; (2) digital nerves are not as apt to be damaged; and (3) accurate reduction and fixation of the osteochondral fracture of the metacarpal head, frequently seen with this dislocation, is possible.


Protection of the myocardium during surgery.

The methods of protection of the myocardium during surgical operations on the heart are described. The relative merits of coronary perfusion and cold arrest of the heart are discussed together with the disadvantages of each technique. The possible place of induced chemical arrest is discussed in relation to coronary perfusion. The technical considerations which affect the choice of method is also considered.


Selective cardiac hypothermia versus coronary perfusion. A study of ischemic complications in two series of 100 consecutive valvular patients.

Myocardial protection, in two parallel series of 100 consecutive valvular patients operated upon between June 1972 and  July 1973 in Broussais Hospital, was afforded in two different ways: one withh hypothermic ischemia (H.I.) as it was advocated by N. Shumway, the other with coronary perfusion (C.P.) of a beating heart with consecutive ischemic periods limited to 20 minutes. Three parameters were used to evaluate the quality of protection: death with primary cardiogenic shock, post-operative myocardial infarction and acute cardiac insufficiency making it necessary to use post-operative inotropic support. Death was 1% (H.I.) and 5% (C.P.). Infarctions were 5% (H.I.) and 8% (C.P.). Acute cardiac insufficiency was: with cardiogenic shock 0% (H.I.) and 4% (C.P.), without cardiogenic shock 8% (H.I.) and 2% (C.P.). Peculiar aspects of myocardial infarction in each series are analyzed, and apparent absence of correlation between aortic cross-clamp time and ischemic complications is discussed. A "myocardial intrinsic factor" seems to be part of each valvular group and appears also to be an important factor in producing ischemic complications.


Ischemic tolerance of the human heart during extracorporeal circulation: clinical experience.

Our clinical experiences with ischemic tolerance of the human heart under the condition of ECC during the last 15 years allow us to state the following points: 1. The healthy human myocardium can be exposed to ischemia without danger on the basis of 15 min and a myocardial temperature of 35 degrees C. This time can be prolonged following the described value of Q10=2. 2. The previous damaged human myocardium is usually not able to cover this range. Frequently ischemic load within these limits causes functional deterioration. So, in such cases ischemic tolerance has to be applied on a distinct lower base. 3. Occasional transgressions of the described ischemic tolerance time without a fatal result do not allow us in our clinic to leave the effective principles in daily routine.


Resistance and tolerance of the myocardium to ischemia.

Since 1967, the authors have abandoned coronary perfusion in valve, particularly aortic, surgery. Some of the difficulties encountered during defibrillation at the period where coronary perfusion was always used have dramatically decreased. Extra-corporeal-circulation is now performed under moderate hypothermia--28 to 30 degrees C--which gives excellent myocardial protection for aortic cross clamping time of 30 to 60 minutes. In coronary artery surgery, the same technique is now used, after having wrongly believed previously that coronary perfusion was indicated. In conclusion, we feel that coronary perfusion is not indicated any more but in very exceptional cases.


Cardioplegic cardiac arrest in aortic surgery.

Cardioplegia according to Bretschneider is able to protect the myocardium during long periods of cardiac arrest and a quiet relaxed heart has in many cases of congenital heart disease been of great surgical advantage. In the last 100 consecutive aortic valve replacements in cardioplegic arrest we have lost 6 patients and none of the deaths seems to be related to the method. In our hands it has been superior to coronary perfusion.


Arteriovenous fistulas of the internal mammary artery. Review of the literature.

Arteriovenous fistulas of the internal mammary artery are rare. This second recorded instance of congestive heart failure from such a fistula occurred in an aged patient 19 years after a gunshot wound. The etiologic and pathophysiologic classifications of 19 cases already reported in the world literature are discussed, as are symptoms, diagnostic methods, and observations during management of this type of fistula.


Effects of left heart bypass on cardiac performance following myocardial ischemia.

A series of nine dogs underwent 20 minutes of myocardial ischemia by cross clamping the aorta while total cardiopulmonary bypass. The four dogs that did not have subsequent left bypass all showed a deterioration of ventricular function curve 30 minutes after restarting the heart beat when compared to their own preischemic values. The five animals which were supported for 30 minutes in left heart after bypass all showed essentially unchanged cardiac function after bypass. This study suggests that an improvement of myocardial performance after ischemic damage can be achieved with left heart bypass.


Ischemia-tolerance following cardioplegic arrest in human patients and in experimental animals.

Cardiac biopsies were taken from the hearts of 40 patients undergoing open heart surgery for acquired and congenital disease. Tissue samples were taken prior to aortic cross-clamping, at the end of the ischemic period, and 20 minutes after reperfusion. Cardiac arrest was induced with Kirsch's cardioplegic solution at mild hypothermia (28-30 degrees C). Electron-microscopy of these tissue samples showed that cardioplegic arrest allows safe recovery with a tolerable degree of cellular alterations following ischemic periods of approximately 45 minutes. This period of reversible ischemia agreed well with similar studies in dogs that were completed by metabolic studies. ATP4-time in these dogs was 60 minutes under conditions closely resembling those at operationmin spite of the similarity between ischemia tolerance times, marked discrepancies existed on the ultrastructural level. Human hearts showed, generally, a much more marked degree of cellular alterations. The good correlation between ultrastructural alterations and the metabolic status of the canine heart can, therefore, not be used to predict levels of metabolites from human electron-micrographies.


Partial cardio-pulmonary bypass in peripheral vascular surgery: case reports.

Two cases of aneurysmatic vascular lesions (a large cranio-encephalic angiodysplasia with arterio-venous communications and a post-traumatic aneurysm of the vertebral artery) are reported; both were of a very difficult surgical approach and were successfully treated with the use of extra-corporeal circulation. The partial cardiopulmonary bypass, with femoro-femoral cannulation, allowed complete recovery of the lost blood, controlled hypotension, and a certain degree of hypothermia. These factors were decisive for the good outcome of the operations.


Visualization of chromatin substructure: upsilon bodies.

Spread chromatin fibers, from isolated eucaryotic nuclei, reveal linear arrays of spherical particles (upsilon bodies), about 70 A in diameter, connected by thin filaments about 15 A wide. These particles have been observed in freshly isolated nuclei from rat thymus, rat liver, and chicken erythrocytes. In addition, upsilon bodies can be visualized in preparations of isolated sheared chromatin, and in chromatin reconstructed from dissociating solvent conditions (i.e., high urea-NaCl concentration). As a criterion for perturbation of native chromatin structure low-angle X-ray diffraction patterns were obtained from nuclear pellets at different stages in the preparation of nuclei fro electron microscopy. These results suggest that the particulate (upsilon body) structures observed by electron microscopy may be closely related to the native configuration of chromatin.


Membrane structure and surface coat of Entamoeba histolytica. Topochemistry and dynamics of the cell surface: cap formation and microexudate.

Treatment of living entamoeba histolytica cells with low concentrations of concanavalin A (con A) and peroxidase results in redistribution of the plasma membrane con A receptors to one pole of the cell where a morphologically distinct region--the uroid--is formed. Capping of con A receptors is not accompanied by parallel accumulation of ruthenium red-stainable components. In capped cells, the pattern of distribution of acidic sites ionized at pH 1.8 (labeled by colloidal iron) at the outer surface and of membrane particles (integral membrane components revealed by freeze-fracture) is not altered over the uroid region. Cytochemistry of substrate-attached microexudate located in regions adjacent to E. histolytica cells demonstrates the presence of con A binding sites and ruthenium red- and alcian blue-stainable components and the absent of colloidal iron binding sites. In a previous report we demonstrated that glycerol-induced aggregation of the plasma membrane particles is accompanied by a discontinuous distribution of colloidal iron binding sites, while con A receptors and acidic sites ionized at pH 4.0 remain uniformly distributed over the cell surface. Taken together, our experiments show that, in E. histolytica cells, peripheral membrane components may move independently of integral components and, also, that certain surface determinants may redistribute independently of others. These results point to the complexity of the membrane structure-cell surface relationship in E. histolytica plasma membranes relative to the membrane of the erythrocyte ghost where integral components (the membrane-intercalated particles) contain all antigens, receptors, and anionic sites labeled so far. We conclude that fluidity of integral membrane components (integral membrane fluidity) cannot be inferred from the demonstration of the mobility of surface components nor, conversely, can the fluidity of peripheral membrane components (peripheral membrane fluidity) be assumed from demonstration of the mobility of integral membrane components.


Selective percutaneous coronary arteriography: an experience with 600 procedures.

Six hundred selective percutaneous coronary angiographies have been performed, using the Judkins technique. The procedure was well tolerated even by critically ill patients. Considering the low risk (mortality: 1.7%), the relatively infrequent complications (2.7%) and the value of the information obtained, selective coronary arteriography is a prerequisite to any meaningful surgery.


Reinitiation of DNA synthesis in senescent human fibroblasts upon fusion with cells of unlimited growth potential.

Postreplicative, "senescent" human fibroblasts were fused to HeLa or to SV-40 transformed human fibroblasts with Sendai virus. DNA synthesis was reinitiated in senescent nuclei in a high proportion of the heterodikaryons. The [3H]thymidine labeling index of senescent fibroblast nuclei in heteropolykaryons was a function of the ratio of HeLa to senescent nuclei.


The site of cellulose synthesis. Hormone treatment alters the intracellular location of alkali-insoluble beta-1,4-glucan (cellulose) synthetase activities.

Membrane preparations from growing regions of 8-day old Pisum sativum epicotyls contain multiple beta-1,4-glucan (cellulose) synthetase activities (UDP- or GDP-glucose: beta-1,4-glucan-glucosyl transferase), and the levels of some of these are influenced by treatments with the growth hormone, indoleacetic acid (IAA). When membranes from control epicotyl segments (zero time) are fractionated by isopycnic sedimentation in sucrose density gradients, all of the synthetase activities are associated mainly with Golgi membrane (density 1.55 g/cm3). After decapitation and treatment of epicotyls with IAA, synthetases also appear in a smooth vesicle fraction (density 1.11 g/cm3) which is rich in endoplasmic reticulum (ER) marker enzyme. Major fractions of these synthetases are not recovered in association with plasma membrane or washed cell walls. When [14-C]sucrose is supplied in vivo to segments +/- IAA, radioactive cellulose is deposited only in the wall. Cellulose or cellodextrin precursors do not accumulate in those membranes in which synthetase activities are recovered in vitro. In experiments where tissue slices containing intact cells are supplied with [14C]sugar nucleotide in vitro, alkali-insoluble beta-1,4-glucan is synthesized (presumably outside the protoplast) at rates which greatly exceeded (20-30 times) those obtained using isolated membrane preparations. Progressive disruption of cell structure results in increasing losses of this high activity. These results are consistent with the interpretation that Golgi and ER-associated synthetases are not themselves loci for cellulose synthesis in vivo, but represent enzymes in transit to sites of action at the wall:protoplast omterface. There they operate only if integrity of cellular organization is maintained.


A disposable venting system.

A disposable venting system that has been utilized in over 300 cardiac operations is described. The system facilitates reduction of bubble formation in the cardiac chambers and removal of air bubbles with minimal time and effort and maximal safety.


Synthesis and turnover of ribulose biphosphate carboxylase and of its subunits during the cell cycle of Chlamydomonas reinhardtii.

The chloroplast enzyme ribulose-1,5-bisphosphate (Ru-1,5-P2) carboxylase (EC 4.1 1.39) is made up ot two nonidentical subunits, one synthesized in the chloroplast and the other outside. Both of these subunits of the assembled enzyme are synthesized in a stepwise manner during the synchronous cell cycle of the green alga Chlamydomonas reinhardtii. The activity of this enzyme increases in the light and this increase is due to de novo protein synthesis as shown by the measurement of the amount of protein and by the pulse incorporation of radioactive arginine in the 18S enzyme peak in linear sucrose density gradients. During the dark phase of the cell cycle, there is little change in the enzymatic activity as well as in the amount of this enzyme. Pulse-labeling studies using radioactive arginine indicated that there is a slow but detectable rate of synthesis of the carboxylase and of its subunits in the dark. Ru-1,5-P2 carboxylase, prelabeled with radioactive arginine throughout the entire light period, shows a similarly slow rate of degradation in the following dark period. This slow turnover of the enzyme in the dark accounts for the steady levels of carboxylase protein and of enzymatic activity during this period. A wide variety of inhibitors of protein synthesis by 70S and 80S ribosomes abolished the incorporation of [3H]arginine into total Ru-1,5-P2 carboxylase during short-term incubation. These results suggest a tight-coordinated control of the biosynthesis of the small and large subunits of the enzyme. This stringent control is further substantiated by the finding that both subunits are synthesized in sychrony with each other, that the ratio of radioactivity of the small to the large subunit remains constant throughout the entire light-dark cycle, and that the rates of synthesis and of degradation of both subunits are similar to that of the assembled enzyme.


Microendocytosis in eosinophilic leukocytes.

Rat peritoneal eosinophils were examined after intraperitoneal infusion either of a mixture of phosphate-buffered saline (PBS) and colloidal gold or of fetal calf serum. These cells characteristically contained vesiculotubular structures, cuplike structures, and small granules during centrifugation. The cup-shaped structures and elaborate labyrinths of vacuole-like spaces increased markedly after injection of the PBS-colloidal gold mixture, presumably as features of heightened microendocytic activity. The vesiculotubular structures increased greatly after infusion of fetal calf serum. A few cyrstalloid granules exhibited fine-structural changes after the PBS-colloidal gold injection, and more numerous crystalloid granules appeared altered after fetal calf serum. Infrequent small granules contained a lucent, crystal-like silhouette after the fetal calf serum injection. Eosinophils evidenced microendocytic uptake of gold spherules into coated vesicles, the cup-shaped structures, and the small granules, but not into the vesiculotubular structures or crystalloid granules after intraperitoneal infusion of the PBS-gold mixture. Strong unmasked acid phosphatase activity in small granules contrasted with the general lack of activity in normal-appearing crystalloid granules and moderate activity in apparently altered crystalloid granules, presumably reflecting active and latent forms of enzyme in the different granules.


Inhibition of interferon secretion by vinblastine.

The plant alkaloids vinblastine and colchicine are known to arrest cells in mitosis by virtue of their binding to spindle protein. These drugs are also capable of binding to microtubule protein and causing these structures to disaggregate into nonfunctional subunits (1, 2). Microtubular structures are thought to be involved in the secretory process of a number of proteins including insulin (7), collagen (4), and thyroid hormone (12). In this report we present our findings on the effects of these two drugs on the synthesis and secretion of interferon in a high producing human foreskin fibroblast strain (FS-4) (11).


Absent right pulmonary artery complicating Tetralogy of Fallot.

A case of tetralogy of Fallot associated with absence of the right pulmonary artery is presented. The patient underwent successful surgical correction. The embryological and clinical aspects of this patient are discussed.


Caerulein secretion by dermal glands in Xenopus laevis.

Since gastrin and its related peptides are secreted by a minority population of widely dispersed cells in mamamalian tissues it has, in the past, been difficult to study the subcellular aspects of their secretion. From published reports (1, 2) it seemed possible that a satisfactory system for such studies might be provided by the skin of certain amphibians such as Xenopus laevis since in these tissues high concentrations of peptides such as caerulein exist, and there is some indication (3) that this, or a similar gastrin-like peptide, may be a dermal gland secretory product. We have therefore explored this possibility by studying the structure, secretory process, and secretory product of the most prominent non mucous type of gland in the skin of X. laevis. These studies clearly demonstrate that most, if not all, of the caerulein in which the skin is contained in secretion granules within the dermal glands and that its release can be specifically evoked by adrenergic stimulation. The release process by a holocrine mechanism expels all of the stored secretion onto the skin surface and thus for biosynthetic studies it should now be possible to synchronize the processes which lead to the replenishment of the peptide.


Bullet embolization to the heart. Report of a case and review of the literature on indications for removal of intracardiac foreign bodies.

The hospital course of a young man with a bullet migration through the venous system to the right ventricle is reviewed. The intraoperative management of foreign bodies to the heart using x-ray image intensifier is employed.


The effects of ionic lanthanum and hypertonic physiological salines on the nervous systems of larval and adult stick insects.

The effects of the electron-opaque tracer ionic lanthanum in various concentrations and of hyperosmotic physiological salines on the nervous system of the stick insect, Carausius morosus, have been studied. Examination of the experimentally treated tissues revealed that the diffusion barrier to the exogenous tracer was maintained in all cases in the adult central and peripheral nervous systems, but not in the hatchling. When hatchling nervous tissues were incubated in 50 mM ionic lanthanum in phyerosmotic physiological saline, the tracer readily infiltrated all the extracellular spaces between axons and glia of all components of the nervous system examined. No difference was noted in this regard between fed and unfed hatchlings, Further, in all cases examined of adults and hatchlings, lanthanum readily surrounded those neurosecretory axons which are found in the neutral lamella, or extracellular nerve sheath, of the insect. The possible meanings of these variations in hatchling and adult nervous systems and in the accessibility of different elements of the nervous system to exogenous ionic lanthanum are discussed.


Ultrastructural changes in chloroplasts resulting from fluctuations in NaCl concentration: freeze-fracture of thylakoid membranes in Dunaliella salina.

The structure of chloroplasts isolated from Dunaliella salina has been studied with respect to changing concentrations of sodium chloride in the culture medium. Freeze-fracture replicas and thin sections of intact chloroplasts do not exhibit any noticeable changes in structure at concentrations ranging between 3.5 and 25% NaCl. Chloroplasts isolated from algal cells that have been acclimatized to the higher salt concentration show a change in the thylakoid membranes. The thylakoid membranes appear compressed over a major portion of the membrane surface, with only the end of the thylakoid membranes unappressed. The number of particles per unit area on the B face is also altered by the salt concentration. The chloroplasts acclimatized to 25% NaCl have about 3 times the number of particles per unit area on a B face of end-membranes as on a comparable face of thylakoid membranes acclimatized to low (3.5% NaCl) salt concentration. These morphological changes can be reversed if the chloroplasts acclimatized to high or low salt concentrations are returned to a medium of different salt concentration prior to freeze-fracturing.


[Atheromatous thromboses of the superficial femoral artery. (Apropos of 400 surgically treated cases)].

The authors report a consecutive series of 400 cases of atheromatous thrombosis of the femoral artery operated on between 1962 and 1972, which gave them an overall idea of the results of surgical treatment with a certain follow-up. This study confirms the importance of the state of the vascular bed beyond the obstruction in the maintenance of permeability after the operation. There are 50 p. 100 long-term failures when the thrombosis extends into the popliteal artery. On the other hand, lesions above the thrombosis, in the aortic or iliac arteries, have little influence on the final result, provided they are treated during the same operation. Lumbar sympathectomy alone gave very poor results but one should note that, in this series, it was reserved for the cases with the poorest prognosis, when any other operation was impossible or contra-indicated. In extreme cases, revascularisation alone of the profunda femoris permitted us in almost all cases to save the limb, leaving intact the symptoms of obstruction of the superficial femoral artery, in particular, intermittent claudication.


[External rotational instability of the knee].

Among cases of recent or long-standing post-traumatic laxity of the knee ligaments, unstable lateral rotation is due to a well-defined anatomical lesion, viz. Rupture of the medial capsular ligament of the knee. This laxity is demonstrated clinically by the existence of an abnormal postero-anterior movement when the tibia is in lateral rotation. Its surgical treatment, suggested by D. Slocum, consists of displacement of the patella tendon. This rupture may exist alone but, usually, is associated with other ligamentous or meniscal lesions for which further specific treatment should be carried out. Slocum's operation has recently been used in the Strasburg Accident Centre. The early results are encouraging.


[Fatigue fractures of the leg bones. Apropos of 19 cases].

The authors report non-traumatic fracture occurring in healthy bond, fatigue fractures of the leg, much less common than fatigue of the metatarsal bones. The authors report here 19 cases, characterised by periosteal apposition and a limited bony fissure which could only be demonstrated towards the second or third week. Its radiological appearance consisted of peri-osteal apposition and bony fissure, and is quite characteristic. However, it is not always obvious and, in some cases, a misdiagnosis of reticulosarcoma was made. It is treated by plaster immobilisation or simple rest and this leads to a cure without sequelae. Progressive and sufficiently prolonged physiotherapy permits recovery of the normal bony structure submitted to new stresses in intensity and duration and should prevent further fractures.


[Post-traumatic fat embolism. Apropos of 272 French cases].

Pulmonary migration of fat globules form a fracture, although anatomically very frequent, rarely gives rise to clinical signs or symptoms. This pathological and clinical entity of fat embolism following trauma, remained for long undiagnosed in France, as the first published cases date from 1949 only; the syndrome is not, however, exceptional for, since 1949, 272 cases have been described including 42 which are published here. Clinical fat embolism affects, above all, adults with fracture of the femoral diaphysis placed in continuous extension. Continuous extension ensures only imperfect immobilisation and permits liberation of fat emboli from the focus whenever the patient moves. Early osteosynthesis is a generally effective means of prevention. The pathogenesis of fat embolism is still debated. It seems that in all cases the clinical signs are due to toxicity of fatty acids liberated from fat emboli. The usual acute neuro-respiratory form occurs after a free interval and associates four groups of symptoms: fever, dyspnea, coma and purpura. There exist forms with mainly neurological or respiratory symptoms, together with mild forms which may raise difficult diagnostic problems. Numerous further examinations have been proposed to permit this diagnosos. One should emphasize, in particular, the fundamental importance of abnormalities of the fundus oculi. The prognosis, which was very poor a few years ago, has been considerably improved with progress in resuscitation based mainly on maintenance of artificial respiration throughout the critical phase which is about 8 to 10 days. Further progress is, however, still possible for there is still a definite mortality.


[Segmental replacement of the prinicipal extra-hepatic bile duct in the dog using a silicone elastomere prosthesis].

The authors present an experimental series of prosthetic replacement of the common bile duct in the dog. The material used was silicone elastomere. In all the dogs studied, a new bile duct became formed around the prosthesis consisting of a fibrous tube covered with mucosa. The follow-up period in the first group of operated dogs was more than 2 1/2 years and in most dogs studied, there was neither biliary cirrhosis nor cholestasis. These encouraging results led the authors to suggest the operation in humans, particularly in carcinoma of the bile ducts.


Chromatography of sterols, alkaloids and other drugs using steam as the mobile phase.

The chromatographic separation of steroids, alkaloids and other drugs with water vapour as the mobile phase has been studied. It is shown that this technique facilitates the analysis and reduces the retention times. The determination of the substances in low concentrations in aqueous solutions or dispersions also appears to be possible.


Determination of 3,3'-dichloro-4,4'-diaminodiphenylmethane in air.

A simple gas chromatographic method is described for the analysis of 3,3'-dichloro-4,4'diaminodiphenylmethane (MOCA) in air. MOCA is quantitatively collected on a Gas-Chrom S tube and the solid sorbent is extracted with acetone. An aliquot sample of the extract is introduced directly into the gas chromatograph, using a microsyringe and the sample is chromatographed on a 1-ft.,silylated 10% Dexsil column. The peak area of MOCA is measured and related to the concentration via a calibration curve prepared with a constant volume of standards. The chloroanilines associated with MOCA cause no interference; however, interference can be expected from any compounds having a retention time identical to that of MOCA. As little as 0.002 mug of MOCA per mul of acetone can be detected by this method.


Open-flask alkaline hydrolysis of amino acid phenylthiohydantoin derivatives.

It is suggested that the open-flask system for the acid hydrolysis of proteins described earlier can also be used to advantage for the alkaline hydrolysis of amino acid phenylthiohydantoin derivatives obtained in protein sequential analysis according to Edman. Results obtained under nitrogen and argon atmospheres are reported for the hydrolysis of 0.5 mumole of each phenylthiohydantoin derivative, which is the amount of protein that is usually employed in an automatic sequenator.


A new approach for obtaining total tryptophan recovery in plasma samples deproteinized with sulphosalicylic acid.

The treatment of plasma with a sodium dodecy sulphate solution before deproteinization with sulphosalicylic acid is described. With this treatment, the liberation of tryptophan from plasma albumin is complete and its determination can be made directly, together with that of other amino acids and related compounds, by automatic ion-exchange chromatography, eliminating the need for an extra analysis for the accurate determination of this amino acid. Results of tryptophan recovery tests, carried out both on plasma samples simply deproteinized with sulphosalicylic acid and on other samples pre-treated with sodium dodecylsulphate solution are reported. These results are also compared with those obtained when trichloroacetic acid was used as deproteinizing agent. A recovery study on a human albumin (fraction V) solution containing a known amount of tryptophan is also described.


Formation of pentafluorophenyldimethylsilylethers and their use in the gas chromatographic analysis of sterols.

The pentafluorophenyldimethylsilyl group is an excellent protecting group for steroid alcohols, giving volatile ethers, detectable at picogram levels in gas chromatography with an electron capture detector. By the use of five reagents, increasingly hindered hydroxyl groups can be protected, so that structural information about unidentified sterols can be obtained. Quantitative derivative formation for a wide range of hydroxyl groups is possible by selection of the correct reagent combination, without affecting unprotected enolizable ketones.


Analysis of Dns-amino acids by liquid chromatography. I. Selection of optimum mobile phase composition for separation of Dns-amino acids on polyvinyl acetate gel.

The separation of a mixture of ten Dns-amino acids (Gns-Gly, -Ala, -Val, -Leu, -Pro, -Hypro, -Met, -Ser, -Asn and -Gln) was carried out by liquid chromatography by using macroreticular polyvinyl acetate gel as a packing material. Different mobile phase systems were investigated, based mainly on mixtures of n-hexane with ethanol, methanol, chloroform, acetone, methyl ethyl ketone, ethyl acetate, dioxane and tetrahydrofuran. The solvent composition was fixed so as to elute all of the components of the sample mixture in a practical period of 2 h. Satisfactory separation of the ten components was obtained with the n-hexane-ethanol (90:10) system. The presence of methanol as a modifier in the n-hexane was effective in reducing the elution time, but the separation was not as satisfactory. Chloroform or dioxane was useful only for the separation of Ser, Asn and Gln. Acetone, methyl ethyl ketone, ethyl acetate and tetrahydrofuran were not suitable for practical separations of Dns-amino acids.


[Estimation of atmospheric benzo(a)pyrene using fluorimetry coupled with liquid scintillation spectrometry (author's transl)].

Methods for the rapid determination of benzo(a)pyrene rest mainly on the separation of the substance by thin-layer chromatography, followed by its spectrophotometric or fluorimetric estimation. Published papers seldom state the recovery results. It is shown here that, with the aid of radioactive benzo(a)pyrene tracer and liquid scintillation spectrometry, recovery becomes poorer and less reproducible as the sample decreases in size (to less than 1 mug benzo(a)pyrene). Coupling fluorimetry with liquid scintillation spectrometry provides an easy means of reducing this common source of error.


Ligand-exchange chromatography of alkaloids.

Alkaloids are separated by liquid chromatography on resins having functional carboxyl groups combined with copper (II) ions. The eluent is aqueous alcohol containing ammonia. Some resins retain alkaloids much better than others. Alkaloids studied included morphine, codeine, strychnine, atropine, papaverine, narcotine, cocaine, quinine, cinchonine and methadone.


Mass fragmentographic determination of 4-hydroxy-3-methoxyphenylglycol (HMPG) in urine, cerebrospinal fluid, plasma and tissues using a deuterium-labelled internal standard.

4-Hydroxy-3-methoxyphenylglycol (HMPG) with the three hydrogen atoms in the side-chain replaced with deuterium (HMPG-D3) was used as the internal standard in the mass fragmentographic determination of free and conjugated HMPG in human urine, cerebrospinal fluid and plasma and in rat urine, liver and brain. HMPG-D3 was added to body fluids or homogenates followed by enzymatic hydrolysis of conjugates. HMPG was extracted with ethyl acetate, converted into the trifluoroacetyl derivative and analyzed by mass fragmentography. HMPG levels were 7.4 nmoles/ml plus or minus 2.8% in human urine, 73 pmoles/ml plus or minus 8.2% in human cerebrospinal fluid, 56 pmoles/ml plus or minus 5.4% in human plasma, 24 nmoles/ml plus or minus 3.6% in rat urine, 0.26 nmoles/g plus or minus 6.2% in rat brain and 99 pmoles/g plus or minus 13% in rat liver. The method is highly specific and sensitive, permitting analysis in small samples or in plasma and in tissues for which previously no methods for HMPG analysis were available.


Quantitation of tryptophan metabolites in rat feces by thin-layer chromatography.

Indole, skatole, indole-3-acetic acid, indole-3-propionic acid, tryptamine, and free tryptophan have been extracted from rat fecal samples and quantitated by thin-layer chromatography. Additional tryptophan metabolites have been recovered in yields of 87-97% from supplemented fecal samples. Detection limits for all the compounds studied were in the 0.1 mug/g feces range. Quantitation was possible in the mug/g range with an estimated accuracy of plus or minus 10%.


Preliminary results on the use of Tenax for the extraction of pesticides and polynuclear aromatic hydrocarbons from surface and drinking waters for analytical purposes.

The use of Tenax GC for the extraction of organic micro-pollutants such as pesticides and polynuclear aromatic hydrocarbons from water has been studied. In the absence of other contaminants and under conditions standardized at the time as optimal, the recovery of such substances is, on average, over 90%, both for the organochlorine and organophosphorus pesticides and for polynuclear aromatic hydrocarbons. Research into the effectiveness of extraction with Tenax from natural waters and water artificially contaminated with surfactants, fats and oils etc., in the laboratory is now being carried out.


Gel filtration of protected peptides on sephadex G-50 in hexamethylphosphoramide containing 5% water.

Gel permeation chromatography on G-50 and G-75 Sephadex gels, using 5% water in hexamethylphosphoramide (phosphoric trisdimethylamide) has been applied to the purification of large protected peptides which are outside the molecular weight range of the Sephadex LH-20-dimethylformamide system.


Determination of disopyramide and its mono-N-dealkylated metabolite in blood serum and urine.

A rapid, precise and accurate assay for disopyramide and mono-N-dealkylated disopyramide concentrations in blood serum and urine is described. The method involves extraction of the drugs from a basic aqueous medium into chloroform, derivatization of the metabolite, purification of the extract and gas chromatographic analysis using an OV-17 liquid phase and flame ionization detection. Unique characteristics of the procedure, direct derivatization in the organic phase and the use of Florisil to separate the drugs from interfering materials, should be applicable to the analysis of other basic drugs in biological specimens.


Determination of serotonin and bufotenin as their dansyl derivatives.

A method is described which permits the determination of serotonin and bufotenin in the same tissue sample. It comprises the following steps: (a) tissue extraction with acetone-0.1 M hydrochloric acid (19:1); (b) reaction of the tissue extract with dansyl (Dns) chloride; (c) pre-separation of O-Dns-bufotenin from O,N-bis-Dns-serotonin and other Dns-amides on a small silica gel column (this step is dispensable if only serotonin or bufotonin is being determined); (d) TLC separation of O-Dns-bufotenin and O,N-bis-Dns-serotonin from other Dns derivatives; (e) quantitative evaluation of the separated compounds by fluorimetry for O-Dns-bufotenin and by fluorimetry or mass spectrometry for the serotonin derivative. The photometer response is linear within the range 0.1-300 nmole. With the mass spectrometric method, 2 pmole of O,N-bis-Dns-serotonin could be determined with a standard deivation of +/-9%. The recovery of the amines from tissue was better than 85%. Reserpine treatment of toads caused a concomitant decrease in serotonin and bufotenin in the brain, but not in the skin of the animals. Repletion of bufotenin in the brain occurs at a higher rate than the repletion of the serotonin pool.


Gas-liquid chromatography and gas-liquid chromatography combined with mass spectrometry of a butyl ester formulation of (2,4-dichlorophenoxy) acetic acid.

A method is presented for the analysis of (2,4-dichlorophenoxy)acetic acid (2,4-D) in commercial formulations, together with concurrent detection and quantitation of related compounds. The method is based on direct gas-liquid chromatographic (GLC) analysis of the formulation to estimate the various esters present, and on an analysis of the total 2,4-D present involving a prior cleanup, alkaline hydrolysis, acidification and n-butylation steps. This procedure also quantitates any other related acids or phenols present. A step which would extract dioxin impurities is also included. The method was checked by the accepted acid-base estimation of 2,4-D content, GLC, and by subjecting radioactive-labelled herbicide to the above procedures.


Separation of fresh tobacco smoke on a packed polar gas chromatographic column prior to on-line analysis by gas chromatography-mass spectrometry using a non-polar capillary column.

Gaseous samples of fresh tobacco smoke were injected on to a packed polar column (2,2-oxydipropionitrile) in all-glass chromatographic system. With the aid of a warm syringe, selected fractions were withdrawn from the bottom of the electron capture detector of the packed column for further injection into an efficient non-polar glass capillary column (SF-96), connectable on-line to a mass spectrometer. The method has permitted the separation and identification of some polar and non-polar components of tobacco smoke, which gave previously mixed, broad, tailing peaks when the smoke was injected directly into a non-polar capillary column.


Analysis of insect pheromones by quadrupole mass fragmentography and high-resolution gas chromatography.

A sensitive technique is described for the analysis of insect pheromones. Insect extracts are separated into fractions by column chromatography and are analyzed directly by quadrupole mass fragmentography on high-resolution glass capillary columns and by mass spectrometry. Applications are illustrated with two important pests. In the codling moth, trans-8, trans-10-dodecadien-1-ol was found to be present at a level of 3.5 ng per female, and in the European grapevine moth trans-7, cis-9-dodecadienyl acetate at a level of 1.6 ng. Both compounds appear to be present in high isomeric purity in the female insect and are absent in the corresponding males.


Gas chromatographic determination of beta-phenethylbiguanide and its metabolite p-hydroxy-beta-phenethylbiguanide in serum and urine.

A gas chromatographic method is presented for the detection of beta-phenethylbiguanide (PEBG) and its metabolite, p-hydroxy-beta-phenethylbiguanide (p-OHPEBG). The procedure is applicable for the determination of the drug and its metabolite in the serum and urine of rats. The detection limit is 0.2 mug PEBG and 0.5 mug p-OH PEBG per ml of serum or urine. A time course study of blood concentration and elimination rate following intraperitoneal injection of 100 mg/kg of PEBG to normal rats was performed. Beta-PEBG was found to be present in the blood and the urine, p-OH PEBG was only detected in the urine. Twenty-four hours following intraperitoneal injection, the urine contained 32% of the administered dose, 20% as unaltered PEBG and 12% as p-OH PEBG.


Organic substances in potable water and in its precursor. III. The closed-loop stripping procedure compared with rapid liquid extraction.

An attempt is made to define the role of gas chromatography in the investigation of organic substances in water, which is important because the handling of water samples before gas chromatographic (GC) analysis depends entirely on the information expected from the subsequent separation, identification and quantification. Practical long-term experience with the previously published closed-loop stripping procedure (with intermediate adsorption on activated carbon) is described and further refinements are reported. A rapid and simple liquid extraction method is described, based on shaking 11 of water with a small volume (0.5-1 ml) of solvent and subsequent high-resolution GC analysis of the extract. Qualitative and semi-quantitative information at the parts per 10(12) level is easily obtained. Further studies of recovery rates under conditions where the volatility and polarity of extracted organic substances are varied are described for both methods. The suitability of both methods for the analysis of different types of water samples is discussed.


Comparative gas chromatographic analysis of narcotics. II. Amphetamine sulphate.

Trace impurities in amphetamine sulphate were studied by a highly sensitive gas chromatographic method. The concentration of these impurities varied considerably between batches whereas the variations within the batches were usually very small. The method is used for the assignment of seizures of amphetamine sulphate to common sources, which in turn may permit chains of illicit distribution of this drug to be traced.


Thin-layer chromatography of hydrazidic monoamine oxidase inhibitor drugs.

Seven solvent systems for the thin-layer chromatographic separation of hydrazidic monoamine oxidase inhibitor drugs have been developed. Detection of compounds at the microgram level was performed with the aid of thirteen spray reagents.


Application of amberlite XAD-2 resin for general toxicological analysis.

The use of Amberlite XAD-2 resin for the extraction of drugs has been extended to the processing of either direct aliquots of blood and stomach contents or aqueous dialysates of the latter, as well as to that of aqueous Stas-Otto deproteinated extracts of tissue. A two-step elution provides the subsequent recovery of standard acid, neutral, basic compounds, and the more polar, water-soluble drugs and drug metabolites. Inorganics may be recovered from the eluate resulting from passage of the original solution through the resin. Extraction with columns of 1-2 g of Amberlite XAD-2 resin is shown to provide the best general approach for separations in pure form of nearly all toxic compounds from up to 500 g of tissue as well as from biological fluids and stomach contents.


Liquid ion exchangers in paper chromatography of steroidal glucosiduronic acids, glucosiduronic esters and free steroids. Influence of concentration of exchanger and counterion.

The chromatographic mobility of steroidal glucosiduronic acids on paper in chloroform-formamide increases as the concentration of ion exchanger in the chloroform phase increases; mobility decreases as the concentration of counterion in formamide increases. Mobility of glucosiduronic esters and of hydroxylated free steroids increases with an increase in concentration of exchanger; small changes in concentration of counterion in the stationary phase do not alter the migration of these nonionizable compounds. Data are presented which suggest that partition of the glucosiduronic acids between the two phases occurs predominantly by an ion-exchange process and that hydrogen bonding plays a secondary role. Partition of the glucosiduronic esters and hydroxylated free steroids appears to occur primarily by a hydrogen-bonding process.


Quantitative determination of epinephrine and norepinephrine in the picogram range by flame ionization gas-liquid chromatography.

A gas-liquid chromatographic method has been developed using the hydrogen flame detector to determine epinephrine (E) and norepinephrine (NE) in blood plasma, red blood cells, serum, and urine. The chromatographic method presents several advantages over other existing techniques. The derivatives enable separation of E and NE and are stable at room temperature with no signs of decomposition. The detection limit for the catecholamines with the hydrogen detector was approximately 0.1 pg. The catecholamines can be determined simultaneously from the same gas-liquid chromatogram. Purification of the catecholamines using the conventional procedure of chromatographing on alumina has been eliminated. With this gas chromatographic method, no by-products are formed that interfere with E and NE determinations. Dopamine, which constitutes the major source of interference in the commonly used fluorometric methods, does not interfere with the E and NE determinations. Norepinephrine and epinephrine values for several physiological fluids are given with the analysis expanded to include red blood cells, the contents of which have not been previously reported.


Use of combined chromatographic methods including thin-layer chromatography for analysis of complex polymer systems. Determination of the polydispersity of block copolymers of styrene and methyl methacrylate by gel permeation, thin-layer and pyrolysis gas chromatography.

A combination of gel permeation chromatography (GPC), thin-layer chromatography (TLC) and pyrolysis gas chromatography (PGC) has been used for investigations of a polymethyl methacrylate-polystyrene-polymethyl methacrylate block copolymer. Continuous distribution of the polymer (40-mg sample) was attained according to the content of the styrene and methyl methacrylate units and of the block copolymer and according to the composition of the copolymer as functions of the hydrodynamic radius of the macromolecules. The polymer was subjected to a preliminary fractionation with an analytical gel chromatograph. The fractions were investigated by TLC, which permitted the separation of the block copolymer and the homopolymers. The composition of the fractions obtained by GPC and TLC was determined by PGC. As a result, it was possible to establish the composition of the block copolymer and its ratio to polymethyl methacrylate in each fraction. This investigation was based on a combination of highly effective fractionation by chromatographic methods with precise quantitative ratios obtained from Benoit's universal calibration graph and from determinations of the composition of the polymer fractions by PGC. The mechanism of the TLC of polymers, including the appearance of artefacts that distort the results of analysis, is also discussed.


Peculiarities in gel permeation chromatography of flexible-chain polymers on macroporous swelling sorbents.

In gel permeation chromatography on macroporous swelling sorbents, deviations from the Benoit principle of universal calibration were observed. It is suggested that these are caused by different degrees of thermodynamic compatibility of the eluted polymers with the sorbent matrix.


Simultaneous determination of acetylmethadol and its active riotransformation products in human biofluids.

A method employing solvent extraction and gas-liquid chromatography has been developed for the quantitative determination of acetylmethadol simultaneously with its two major biotransformation products, noracetylmethadol and dinoracetylmethadol. Noracetylmethadol and dinoracetylmethadol are analyzed following their conversion to the corresponding amides. The amide structure is confirmed by the use of chemical ionization mass spectroscopy and infrared spectroscopy. The method can be used to determine the concentration of acetylmethadol and these compounds in plasma samples from acetylmethadol maintenance subjects. Methadol and normethadol do not attain neasurable plasma levels. Urine contains predominantly noracetylmethadol and dinoracetylmethadol. Evidence was also obtained for the urinary excretion of acetylmethadol, methadol and normethadol. A mean quantity equal to 28% of the administered dose was excreted in the urine of a 48-h dosing interval as acetylmethadol and metabolites.


Determination of perphenazine and its sulphoxide metabolite in human plasma after therapeutic doses by gas chromatography.

A gas chromatographic method for the determination of perphenazine (Tri-lafon) and its main metabolite in human plasma, perphenazine sulphoxide, has been developed. The procedure involves the use of an electron capture detector and permits the determination of the drug and its metabolite at concentrations down to 0.2 mug/l. This is sufficient for analyzing plasma from patients on ordinary treatment with perphenazine. Tests for specificity revealed no interference by nortriptyline or biperidine.


Dual column gas chromatographic system for use in mass spectral determination of nitrosamines.

A packed gas chromatographic column and a support coated open tubular (SCOT) column are connected in series. Between the columns are two micro-volume switching valves, one enabling solvent to be vented. Short retention nitrosamines are passed through both columns, whereas longer retention nitrosamines by-pass the SCOT column by means of the other switching valve.


Trace analysis of volatile N-nitroso compounds by combined gas chromatography and thermal energy analysis.

Thermal energy analysis (TEA) has been combined with gas chromatography (GC). The new GC-TEA technique is highly specific to compounds which contain heat labile nitrosyl groups. Because of the specificity of the technique, full use may be made of the TEA sensitivity. Analysis by direct injection of solutions containing less than 1 ng/ml N-nitroso compound is demonstrated.


Separation of resin acids from fatty acids in relation to environmental studies.

Kraft mills pulping coniferous wood species discharge to their receiving waters effluents containing mixed resin and fatty acids. To study the fish-toxic properties of these fractions it is desirable to be able to separate them without causing double bond isomerizations in the acid-labile resin acid fraction. A method is described using toluenesulphonic acid as catalyst under very mild conditions and four examples of its application are given. It was not possible to separate tall oil samples without a low degree of resin acid isomerization.


[Separation of herbicides by high-performance liquid chromatography. Influence of water (author's transl)].

The influence of the amount of water dissolved in dichloromethane on the chromatographic separation of herbicides was studied. The selectivity of the mobile phase was demonstrated and compared to another system, dichloromethane modified with 2-propanol. The high efficiency of microporous packing was also demonstrated.


Automated chromatographic determination of chlorhexidine in pharmaceutical preparations.

An automated method for the determination of chlorhexidine (Hibitane) and its salts in formulated pharmaceutical products is described. The equipment consists of a high efficiency liquid chromatograph, a variable wavelength high sensitivity ultraviolet spectrophotometric detector, the output of which is monitored simultaneously by a suitable recorder, and a digital computer. The sample is automatically introduced onto a 10-cm silica gel column by use of a slide valve. Results are calculated and printed out by the computer.


Application of densitometry to the qualitative and quantitative evaluation of pharmaceutical colourants.

Samples from all the British manufacturers of food grade Amaranth and Sunset Yellow FCF were examined by a densitometric thin-layer chromatographic procedure which provides a rapid and convenient method for producing a qualitative and quantitative statement on the impurity profiles of dye materials. Wide variations in the impurities were found which could account for variations in toxicological results. The method could be incorporated into official specifications to control more precisely the quality of food grade dyes.


Determination of a new cerebral vasodilator 2,6-dimethyl-4-(3-nitrophenyl)-1,4-dihydropyridine-3,5-dicarboxylic acid 3-[2-(N-benzyl-N-methylamino)]-ethyl ester 5-methyl ester hydrochloride (YC-93) in plasma by electron capture gas chromatography.

A highly sensitive method for the quantitative determination of 2,6-dimethyl-4-(3-nitrophenyl)-1,4-dihydropyridine-3,5-dicarboxylic acid 3-[2-(N-benzyl-N-methylamino)]-ethyl ester 5-methyl ester hydrochloride (YC-93) in plasma is described. After extraction, YC-93 was oxidized to a pyridine analogue with nitrous acid and detected by electron capture gas chromatography. The sensitivity was 2-3 ng/ml, which is sufficient to determine plasma concentrations of YC-93 after oral administration of clinical doses to humans.


Determination of pinazepam and its metabolites in serum, urine and brain by gas-liquid chromatography and mass spectrometry.

A sensitive and specific assay, involving electron capture gas-liquid chromatography, has been developed for the identification of pinazepam and its metabolites in serum, urine and brain samples from dogs and rats after single or repeated oral administration of the drug. Serum and urine samples from healthy humans after a single oral administration have also been analysed. The identity of gas-liquid chromatographic peaks has been established by mass spectrometry. In blood serum and brain, only pinazepam and its N-depropargylated product (demethyldiazepam) were found; from urine, 3-hydroxypinazepam and oxazepam were also recovered. The sensitivity of the gas-liquid chromatographic method is of the order of 5-10 ng of pinazepam and 15-20 ng of the other three benzodiazepines per ml of serum.


Non-specific binding in the affinity chromatography of chymotrypsin.

The strong and specific binding of chymotrypsin on chromatographic columns containing agarose substituted with N-sigma-amino caproyl-D-tryptophan methyl ester is abolished when the sigma-amino groups on the surface of the enzyme are reacted with acetic anhydride. Because the catalytic properties of the acetylated chymotrypsin are identical to those of the underivatized enzyme, it is concluded that the high affinity of chymotrypsin for this column is not due solely to biospecific inhibitor binding, which is by itself very weak, but requires reinforcement through weak non-specific interactions with the column support. It is postulated that these non-specific interactions include electrostatic interactions between agarose matrix and positively charged lysyl residues on the enzyme. The results demonstrate for the first time that residues on the surface of an enzyme not associated with its active site can play an important role in some chromatographic systems previously thought to be based on purely biospecific interactions.


Estimation of urinary diamines and polyamines by thin-layer chromatography.

The dansylated derivatives of ammonia, 1,3-diaminopropane, 1,4-diaminobutane, 1,5-diaminopentane, spermidine, histamine, and spermine were separated by one ascending development in chloroform-triethylamine (5:1) on a 250-mum silica gel 60 plate. Putrescine, cadaverine, spermidine, and spermine in human urine were quantitated by a direct scan of the fluorescent intensity of the spots corresponding to these compounds. Higher amounts of spermidine and spermine were found in the urines of cancer patients compared to the values of these substances in normal urine.


Methods of identification and confirmation of abusive drugs in human urine.

A thin-layer chromatography (TLC) procedure is described to be used as the initial drug detection method for urine surveillance in a drug abuse treatment program. While the TLC method is sufficiently sensitive, it is prone to false-positive results. For this reason, two other drug detection methods (gas-liquid chromatography and radioimmunoassay) have been incorporated to confirm positive results obtained with TLC. The combined methodologies result in a urine surveillance procedure that is versatile, sensitive and highly reliable.


[Investigations on the production of mycotoxins and their quantitative evaluation. I. The production of penicillic acid by Penicillium cyclopium (author's transl)].

A convenient thin-layer chromatographic screening procedure for the analysis of mycotoxins produced by Penicillium cyclopium is described. The production of penicillic acid is followed during a growth period of 44 days at two different temperatures. Quantitative evaluation is performed by UV densitometry at 234 nm. A biological profile is recorded showing the different ratios of four metabolites produced by Penicillium cyclopium and the definite effect of the growth temperature on the formation of penicillic acid in relation to the other metabolites.


High-performance liquid chromatographic determinaton of pentaerythritol in plasma.

A sensitive analysis of pentaerythritol in plasma has been devised, based on the formation of its tetra-p-methoxybenzoate derivative and high-performance liquid chromatography employing an ultraviolet photometric detector. The method permits analysis of pentaerythritol in the ppm range.


0-(2,3,4,5,6-pentafluorobenzyl)hydroxylamine hydrochloride as a sensitive derivatizing agent for the electron capture gas liquid chromatographic analysis of keto steroids.

0-(2,3,4,5,6-pentafluorobenzyl)hydroxylamine hydrochloride was used to prepare oximes of steroids with keto groups in selected positions; 3,17 and 20-monoketo; 3,17 and 3,20-diketo. Some of the 3-keto steroids had hindered 17-hydroxyl groups which were not readily amenable to esterification with perfluoroanhydrides, the most commonly used derivatizing agents for electron capture gas chromatographic analysis of hydroxy steroids. The oximes were readily prepared from 5 ng of each of the compounds tested, and with testosterone it was demonstrated that the derivative could be prepared from as little as 0.1 ng. The derivatives were stable to gas chromatography and extremely sensitive to electron capture detection. The sensitivity ranged from 1.5 X 10(4) coulombs per mole of progesterone. Because of the ease of preparation of the derivatives, their stability in common solvents and analytical manipulative techniques, the reagent would be suitable for the micro analysis of biologically significant keto steroids by electron capture gas chromatography.


Improved resolution of natural diacylglycerols by gas-liquid chromatography on polar siloxanes.

A new cyanoalkylphenylsiloxane (SILAR 5CP) liquid phase is shown to possess sufficient polarity to permit improved GC separations of natural diacylglycerols based on unsaturation and positional placement of fatty acids as well as on molecular weight, which was previously possible only on ethylene glycol succinate polyesters. Unlike the polyesters, the polar siloxane polymer has moderate thermal stability and provides GC columns which can be used for several months without replacing the packing. The GC analyses were made with conventional columns containing 3% SILAR 5CP on Gas Chrom Q at 270 degrees C isothermally. The diacylglycerols were chromatographed as the TMS ethers. Excellent seperations were obtained for the 1,2(2,3)- and 1,3-diacyglycerols derived from corn, linseed, peanut and cod liver oils and for the 1,2-diacyl-sn-glycerols from hepatic glycerophospholipids.


Systems for automated multiple pesticide residue analysis.

This article reviews the development and current status of automated multiple pesticide residue analysis. Various configurations of equipment that perform specific steps in a method of analysis are described. These include the use of Technicon modules, automatic injectors for gas chromatography, specially designed units and data processing systems.


Microanalytical methodology relating to the identification of insect sex pheromones and related behavior-control chemicals.

Methods currently in use in the extraction, isolation, and identification of insect sex attractants, which are needed for use in pest control, are described briefly. Because amounts of these materials in insects are generally minute, specialized microanalytical methods and techniques have been developed to aid in the determination of their chemical structures.


A survey of the chromatographic analysis of natural insect juvenile hormones and the insect growth regulator, altosid.

A brief survey of gas and liquid chromatographic methods for analysis, purification and quantitation of natural insect juvenile hormones and one commercially used analoh, altosid (methoprene) insect growth regulator, includes references to the most recent literature.


Pituitary TSH in normal subjects and in patients with asymptomatic atrophic thyroiditis: Evidence for its immunological heterogeneity.

Biological and radioimmunological measurements of pituitary TSH concentration were performed in 22 cases of asymptomatic atrophic thyroiditis and in 18 controls. Whilst bioassay revealed the presence of a greater pituitary TSH concentration in thyroiditis cases, radioimmunoassay failed to confirm such a difference. The reason therefore seems to lie in the presence in thyroiditis pituitatries of a TSH which reacted in the bioassay but showed only a weak affinity for the anti-hTSH antiserum. The slopes of radioimmunological dilution curves of the pituitary extracts were indeed significantly lower with thyroiditis pituitaries than with controls. When the whole population sample was considered, a negative correlation existed between the ratio of biological and radioimmunological TSH determinations (B/I) and the slope of the corresponding dilution curves. Since in radioimmunoassay a low displacement slope is indicative of a weak immunological affinity of the antigen for the antiserum, this demonstrated negative correlation suggests together with a high B/I ratio, even in normal people.


Preovulatory progestins: Observation on their source in rhesus monkeys.

Progesterone (P) and 20alpha-hydroxypregen-4-en-3-one (20alpha-ol)concentrations were measured in systemic and ovarian vein plasma of adult cycling female rehsus monkeys by gas liquid chromatography with electron capture detection. Blood samples and ovaries were removed at various times during the intermenstrual period. The animals were then reclassified according to the stage of the ovarian cycle and the morphology of the ovary. Very little P or 20alpha-ol is secreted by the ovary during the follicular phase until the largest developing follicle measures 5.6 mm in diameter. In the prevular period, the concentrations of both P and 20alpha-ol in the two ovarian veins are nearly equal. As the luteal phase develops, the ratios of progestin concentrations in the ovarian vein change, more progestin being found in ovarian venous plasma of the ovary with the corpus luteum. The secretion of preovulatory progestin can be correlated with large amounts of interstitial gland cells that appear before ovulation in the ovary. In a second experiment, follicular development was estimated by a rapid radioimmunoassay for estrogen in the systemic circulation. When the systemic plasma levels of immunoreactive estrogen exceeded 150 pg/ml, ovarian venous samples of blood were collected from each ovary and analyzed by radioimmunoassay for estradiol-17beta and progesterone. Ffteen min after removal of the ovary that contained the mature Graafian follicle, blood was drawn from the contralateral ovarian vein. In this preovulatory condition, the progesterone concentrations in both ovarian veins greatly exceeded those in the systemic circulation. In 2 animals, removal of the ovary that contained the mature Graafian follicle did not lower the blood levels of progesterone in the contralateral ovarian vein after 15 min. These data suggest that both ovaries of the rhesus monkey secrete preovulatory progestin which is available for physiological action.


Dissociated responses of plasma testosterone and estradiol to human chorionic gonadotropin in adult men.

Plasma testosterone (T), dihydrotestosterone (DHT) and estradiol (E2) were determined by radioimmunoassay in 10 normal males receiving hCG im 5000 IU on days 1, 2 and 3. The mean increase of plasma steroid on days 2, 3 and 1, respectively, was: 1.42, 1.79 and 1.87 times for T; 1.17, 1.56 and 1.49 times for DHT; 4.04, 3.29 and 2.33 times for E2. While T was still significantly increasing from day 2 to day 4, E2 significantly decreased. A positive correlation (P less than 0.01) was found between the basal T and the E2 peak after hCG, suggesting a release of E2 from a storage compartment in the testis. No significant change of either steroid was detected 4 h after the first hCG injection. In 6 cases of primary male hypogonadism, the mean basal values of T to hCG was defective, despite considerable individual variations. In 14 males with gonadotropin deficiency, basal values of T and E2 were very low; the T response to hCG ranged from undetectable to dramatic, and was correlated with the degree and duration of previous exposure to gonadotropin; and impaired response of E2 in all cases porvides a better estimate of the actual gonadotropin deficiency.


Elevated prostaglandins and suppressed parathyroid hormone associated with hypercalcemia and renal cell carcinoma.

A patient with renal cell carcinoma and hypercalcemia was found to have suppressed serum parathyroid hormone levels but striking elevations of immunoreactive prostaglandins in plasma and metastatic tissue. It was hypothesized that prostaglandins may have either played a role in the hypercalcemia or were part of a counterregulatory event.


Insulin responses to crude cholecystokinin-pancreozymin in normal subjects, in patients with chronic pancreatitis and patients with mild maturity onset diabetes.

The effects of repeated injections of 75 U crude cholecystolinin-pancreozymin (CCK-PZ) at increasing plateau glucose concentrations achieved by glucose infusion were studied in 15 controls, 8 chronic pancreatitics and 8 mild maturity onset diabetics. In control subjects CCK-PZ alone caused minor insulin release but proportinally greater secretion with increasing blood glucose concentrations. Chronic pancreatitis patients who had normal responses to intravenous glucose responded normally to the CCK-PZ but at significantly higher plateau glucose levels. Diabetics had no response to IV glucose boluses of 5 g or 10 g, but with glucose infusions of 250-500 mg/min had almost normal insulin responses to CCK-PZ. The responses to CCK-PZ plus glucose were greater than either stimulus alone, indicating an interaction between these and the beta cell. These studies suggest that the gut homone-receptor in the beta cell is intact in maturity onset diabetes and chronic pancreatitis, whether the glucose receptor is normal or defective. The peptide-responsible in the crude CCK-PZ is not secretin, glucagon or gut glucagon, but may be gastric inhibitory polypeptide (GIP) since pure CCK-PZ has no insuli releasing properties.


The effect of intravenous prostaglandin F2alpha on serum luteinizing hormone, follicle-stimulating hormone and plasma cortisol in normal men.

Prostaglandin F2alpah (PGF2alpha) was given intravenously in a maximun dose of 24 mug per min to 6 normal male volunteers. There was no change in serum luteinizing hormone, follicle-stimulating hormone, or blood glucose. Plasma cortisol fluctrated but no substained rise was demonstrated.


Glucagon stimulation of plasma renin activity in humans.

Administration of 0.2 mg of glucagon by intravenous bolus resulted in an increase in plasma renin activity (PRA) in 2 of 5 normal volunteers on their usual diet. Two of the nonresponders subsequently showed a PRA response to glucagon after sodium depletion. A lower dose of glucagon (0.01 mg) had no effect on PRA despite a 31 mg/100 ml rise in blood glucose and peak plasma glucagon of over 2000 pg/ml. In conclusion, glucagon can stimulate PRA but it is not a potent stimulator; its effect may be potentiated by sodium depletion.


Diversion of peripheral thyroxine metabolism from activating to inactivating pathways during complete fasting.

In 9 euthyroid obese volunteers, as previously reported, 4 weeks of total caloric deprivation resulted in a striking decrease in serum 3,5;3'-triiodothyronine (T3) concentration. The present studies reveal that this decrease in serum T3 is accompanied by a proportionately similar increase in the serum concentration of 3,3',5' -T3 (reverse T3; rT3). In four additional obese volunteers given suppressive doses of sodium-Lthyroxine (T4) for 1 month prior to fasting, serum T3 concentration declined sharply during a 6-11 day period of fast, while rT3 concentration increased strikingly. Concentrations of both T3 and rT3 returned to control values during a 5 day period of refeeding. The findings indicate that caloric deprivation results in an alteration in peripheral T4 metabolism away from generation of T3 and toward the generation of rT3. Since the former is more active than T4, and the latter is essentially inactive, caloric deprivation appears to shunt peripheral T4 metabolism from activating to inactivating pathways.


Thyroid-stimulating hormone levels in idiopathic euthyroid goiter.

Utilizing a double antibody radioimmunoassay for human TSH we compared distribution of serum TSH in 167 normal individuals and 51 patients with idiopathic euthyroid goiter. In addition to being clinically euthyroid both groups had normal total thyroxine, and free thyroid index. Forty-two percent of the goiter group had high TSH and the general distribution of TSH values in the goiterous patients was significantly higher than normal (P less than 0.001). Anlysis of subgroups of the normal and goiter populations indicated that the high TSH could not be attributed to age, sex, use of birth control pills or differences between diffuse and multinodular goiter. TSH levels were significantly higher in patients with goiter less than 1 yr compared to goiter greater than 1 yr (P less than 0.025). In patients with goiter greater than 1 yr the TSH levels remained significantly higher than normal (P less than 0.025). These results support the hypothesis that TSH plays a role in the genesis of idiopathic goiter. The elevation may be present only early in the course of goiter development but is also present in a significant number of patients with long standing goiter.