Renaturation of formyltetrahydrofolate synthetase from urea and guanidinium chloride solutions

Formyltetrahydrofolate synthetase from Clostridium cylindrosporum and Clostridium acidi-urici was denatured in 6 M urea and 4 M guanidinium chloride. Viscometric, fluorimetric and ultracentrifugal measurements were used to determine that the protein is completely unfolded under these conditions. The polypeptide chains refold upon dilution of the denaturant-protein solutions to give final concentrations of 0.5 M urea or 0.1 M guanidinium chloride. In the presence of NH4+, but not in its absence, the refolded proteins associate to produce the catalytically active tetramer. Refolding and reassociation were followed by measuring changes in protein fluorescence and by determination of sedimentation constants. Under most conditions 80% of the enzymic activity is recovered.     

Inactivation of pyridoxal phosphate dependent enzymes by mono- and polyhaloalanines

beta,beta-Dichloro- and beta,beta,beta-trifluoroalanine irreversibly inactivate a number of pyridoxal phosphate dependent enzymes which catalyze beta- or gamma-elimination reactions. The inactivation is time dependent and the rate of inactivation is first order in enzyme concentration. This suggests that inactivation is due to covalent modification of the enzyme by a species generated at the active site from the polyhaloalanine (i.e., suicide inactivation). Monohaloalanines are substrates and do not inactivate. For gamma-cystathionase, covalent and stoichiometric attachment of [1-14C]beta,beta,beta-trifluoroalanine was shown. It is proposed that the mechanism of inactivation involves Schiff base formation between inactivator and enzyme-bound pyridoxal and subsequent elimination of HC1 from dichloroalanine or HF from trifluoroalanine. This results in the formation of a beta-halo-alpha,beta unsaturated imine, an activated Michael acceptor. Michael addition of a nucleophile at the active site leads to covalent labeling of the enzyme and inactivation. Alanine racemase is also inactivated by the two polyhaloalanines. Glutamate-pyruvate and gultamate-oxaloacetate transaminase are inactivated by monohaloalanines but not by polyhaloalanines.     

Bilateral pulmonary hilar lymphadenopathy. An unusual manifestation of metastatic renal cell carcinoma

The significance of glucagon for the alterations in carbohydrate and fat metabolism during swimming has been evaluated. Fed, male rats were used. Blood was drawn by cardiac puncture for glucose analysis and either rabbit-antiglucagonserum (A-rats) or normal rabbitserum (N-rats) injected. Twenty-nine rats were then forced to swim (S-rats) with a tail weight for 60 min, while 16 rats were resting controls (C-rats). Subsequently blood was drawn and samples of liver and muscle tissue collected. In SN-rats glucagon concentrations increased from 152 +/- 18 (S.E.) pg/ml (CN-rats) to 332 +/- 61 (P less than 0.05), while liver glycogen decreased (P less than 0.001) and blood glucose increased (P less than 0.05). In SA-rats, however, the changes in liver glycogen and blood glucose were halved indicating that increased glucagon secretion enhances hepatic glycogen depletion during prolonged exercise. NEFA rose in SA-rats (P less than 0.005) as well as in SN-rats (P less than 0.05). Glycerol concentrations, however, only increased in SA-rats (P less than 0.05) indicating a shift towards lipid combustion in antibody treated rats. Muscle glycogen and plasma insulin diminished and blood lactate increased uniformly in exercised rats.     

Immunoreactive growth hormone levels in pigs fed protein or energy restricted diets during the postweaning period

Developmental changes in growth hormone levels during protein or energy malnutrition was studied in weaned pigs. Three or 4 weeks old pigs from control dams were fed a control diet (18% protein), an energy-restricted diet (18% protein) or a low protein diet (6% protein) for 8 weeks. Energy restriction was achieved by feeding the control diet in amounts that allowed very little growth. After the restriction period, all pigs were fed the control diet ad libitum for another 8 weeks. Blood samples were collected at intervals throughout the experiment and the plasma was analyzed for growth hormone by radioimmunoassay. Post weaning protein deprivation resulted in higher growth hormone levels during the restriction period as compared to control pigs or pigs with a restricted energy intake.     

Electrophysiologic effects of propranolol on sinus node function in patients with sinus node dysfunction

The electrophysiologic effects of intravenously administered propranolol (0.1 mg/kg) on three parameters of sinus node function were examined in ten symptomatic patients with sinus node dysfunction. The patients ranged in age from 26 to 79 years. Symptoms ranged from fatigue to frank syncope. Sinoatrial (SA) block and sinus pauses were observed in one patient; sinus pauses alone were observed in three patients. Five (5/10) patients had intraatrial block; three (3/10) patients had atrioventricular block; four (4/10) patients had an intraventricular conduction disturbance. At the time of electrophysiologic study, two patients had a control spontaneous sinus cycle length that exceeded 1000 msec. Following propranolol, the mean spontaneous cycle length increased by 17.4% (924 to 1085 msec, P less than 0.005) and spontaneous second degree SA block reappeared in the one patient. The maximum escape cycle ranged from 116% to 229% of the prepacing spontaneous cycle length and was considered to be prolonged in two of ten patients. Propranolol had no significant effect on the maximum escape cycle/prepacing cycle length X 100 (%). The estimated sinoatrial conduction time (SACT) was determined in seven patients and ranged in value from 120 to 238 mes. Propranolol increased the mean value of the estimated SACT from 179 to 213 msec, P less than 0.025. Propranolol may cause marked bradyarrhythmias in some patients with sinus node dysfunction, and should be used with caution in these patients.     

Persistent obstructive jaundice, cholangitis, and biliary cirrhosis due to common bile duct stenosis in chronic pancreatitis

Long strictures of the intrapancreatic portion of the common bile duct were found in 6 patients with chronic pancreatitis. These strictures were responsible for painless obstructive jaundice, recurrent cholangitis, secondary biliary cirrhosis, and chronic abdominal pain difficult to distinguish from that caused by pancreatitis. Endoscopic retrograde cholangiopancreatography and intraoperative cholangiography were invaluable in making the diagnosis and in planning surgical correction. Decompression of the biliary tree by anastomosis of the gallbladder or common duct to the small intestine completely relieved symptoms and allowed liver function to improve significantly. Common duct stricture as a complication of chronic pancreatitis should be considered in the differential diagnosis of extrahepatic biliary obstruction and whenever surgical treatment of chronic pancreatitis is contemplated.     

Innovations in beef production systems that enhance the nutritional and health value of beef lipids and their relationship with meat quality

Consumers are becoming more aware of the relationships between diet and health and this has increased consumer interest in the nutritional value of foods. This is impacting on the demand for foods which contain functional components that play important roles in health maintenance and disease prevention. For beef, much attention has been given to lipids. This paper reviews strategies for increasing the content of beneficial omega-3 polyunsaturated fatty acids (PUFA) and conjugated linoleic acid (CLA) and reducing saturated fatty acids (SFA) in beef. Particular attention is given to intramuscular fat (IMF) and the relationships between fatty acid composition and key meat quality parameters including colour shelf life and sensory attributes. Despite the high levels of ruminal biohydrogenation of dietary PUFA, nutrition is the major route for increasing the content of beneficial fatty acids in beef. Feeding grass or concentrates containing linseed (rich in α-linolenic acid, 18:3n-3) in the diet increases the content of 18:3n-3 and its longer chain derivative eicosapentaenoic acid (EPA, 20:5n-3) in beef muscle and adipose tissue, resulting in a lower n-6:n-3 ratio. Grass feeding also increases docasahexaenoic acid (DHA, 22:6n-3). Feeding PUFA rich lipids which are protected from ruminal biohydrogenation result in further enhancement of the PUFA in meat with concomitant beneficial improvements in the ratio of polyunsaturated:saturated fatty acids (P:S ratio) and n-6:n-3 ratio. The main CLA isomer in beef is CLA cis-9, trans-11 and it is mainly associated with the triacylglycerol lipid fraction and therefore is positively correlated with level of fatness. The level of CLA cis-9, trans-11 in beef is related to (1) the amount of this isomer produced in the rumen and (2) synthesis in the tissue, by delta-9 desaturase, from ruminally produced trans vaccenic acid (18:1 trans-11; TVA). Feeding PUFA-rich diets increases the content of CLA cis-9, trans-11 in beef. Trans-fatty acids in foods are of rising importance and knowledge of the differential effects of the individual trans isomers is increasing. TVA is the major trans 18:1 isomer in beef and as the precursor for tissue CLA in both animals and man should be considered as a neutral or beneficial trans-isomer. Increasing the content of n-3 PUFA in beef can influence colour shelf life and sensory attributes of the meat. As the content of n-3 PUFA increases then sensory attributes such as "greasy" and "fishy" score higher and colour shelf life may be reduced. Under these situations, high levels of vitamin E are necessary to help stabilise the effects of incorporating high levels of long chain PUFA into meat. However, grass feeding not only increases n-3 PUFA and CLA but, due to its high content of vitamin E, colour shelf life is improved. It is evident that opportunities exist to enhance the content of health promoting fatty acids in beef and beef products offering opportunities to add value and contribute to market differentiation. However, it is imperative that these approaches to deliver "functional" attributes do not compromise on the health value (lipoperoxidation) or the taste of beef products.     

Fat deposition, fatty acid composition and meat quality: A review

This paper reviews the factors affecting the fatty acid composition of adipose tissue and muscle in pigs, sheep and cattle and shows that a major factor is the total amount of fat. The effects of fatty acid composition on meat quality are also reviewed. Pigs have high levels of polyunsaturated fatty acids (PUFA), including the long chain (C20-22) PUFA in adipose tissue and muscle. The full range of PUFA are also found in sheep adipose tissue and muscle whereas cattle ‘conserve’ long chain PUFA in muscle phospholipid. Linoleic acid (18:2n − 6) is a major ingredient of feeds for all species. Its incorporation into adipose tissue and muscle in relation to the amount in the diet is greater than for other fatty acids. It is deposited in muscle phospholipid at a high level where it and its long chain products eg aracidonic acid (20:4n − 6) compete well for insertion into phospholipid molecules. Its proportion in pig adipose tissue declines as fat deposition proceeds and is an index of fatness. The same inverse relationships are not seen in ruminant adipose tissue but in all species the proportion of 18:2n − 6 declines in muscle as fat deposition increases. The main reason is that phospholipid, where 18:2n − 6 is located, declines as a proportion of muscle lipid and the proportion of neutral lipid, with its higher content of saturated and monounsaturated fatty acids, increases. Oleic acid (18:1cis − 9), formed from stearic acid (18:0) by the enzyme stearoyl Co-A desaturase, is a major component of neutral lipid and in ruminants the same enzyme forms conjugated linoleic acid (CLA), an important nutrient in human nutrition. Like 18:2n − 6, -linolenic acid (18:3n − 3) is an essential fatty acid and is important to ruminants since it is the major fatty acid in grass. However it does not compete well for insertion into phospholipid compared with 18:2n − 6 and its incorporation into adipose tissue and muscle is less efficient. Greater biohydrogenation of 18:3n − 3 and a long rumen transit time for forage diets also limits the amount available for tissue uptake compared with 18:2n − 6 from concentrate diets. A positive feature of grass feeding is that levels of the nutritionally important long chain n − 3 PUFA are increased ie EPA (20:5n − 3) and DHA (22:6n − 3). Future research should focus on increasing n − 3 PUFA proportions in lean carcasses and the use of biodiverse pastures and conservation processes which retain the benefits of fresh leafy grass offer opportunities to achieve this. The varying fatty acid compositions of adipose tissue and muscle have profound effects on meat quality. Fatty acid composition determines the firmness/oiliness of adipose tissue and the oxidative stability of muscle, which in turn affects flavour and muscle colour. Vitamin E is an essential nutrient, which stabilises PUFA and has a central role in meat quality, particularly in ruminants.     

Occurrence, synthesis, and mammalian cell cytotoxicity and genotoxicity of haloacetamides: an emerging class of nitrogenous drinking water disinfection byproducts

The haloacetamides, a class of emerging nitrogenous drinking water disinfection byproduct (DBPs), were analyzed for their chronic cytotoxicity and for the induction of genomic DNA damage in Chinese hamster ovary cells. The rank order for cytotoxicity of 13 haloacetamides was DIAcAm > IAcAm > BAcAm > TBAcAm > BIAcAm > DBCAcAm > CIAcAm > BDCAcAm > DBAcAm > BCAcAm > CAcAm > DCAcAm > TCAcAm. The rank order of their genotoxicity was TBAcAm > DIAcAm approximately equal to IAcAm > BAcAm > DBCAcAm > BIAcAm > BDCAcAm > CIAcAm > BCAcAm > DBAcAm > CAcAm > TCAcAm. DCAcAm was not genotoxic. Cytotoxicity and genotoxicity were primarily determined by the leaving tendency of the halogens and followed the order I > Br > > Cl. With the exception of brominated trihaloacetamides, most of the toxicity rank order was consistent with structure-activity relationship expectations. For di- and trihaloacetamides, the presence of at least one good leaving halogen group (I or Br but not Cl) appears to be critical for significant toxic activity. Log P was not a factor for monohaloacetamides but may play a role in the genotoxicity of trihaloacetamides and possible activation of dihaloacetamides by intracellular GSH and -SH compounds.