bluuebunny/crossref_metadata_2025 huggingface dataset:
Inactivation of Escherichia coli with Power Ultrasound in Apple Cider
Authors: Ugarte‐Romero Edgar , Feng Hao , Martin Scott E. , Cadwallader Keith R. , Robinson Scott J.
Published: August 2010

The use of acoustic energy to secure apple cider safety was explored. Inactivation tests were performed with Escherichia coli K12 at 40 °C, 45 °C, 50 °C, 55 °C, and 60 °C with and without ultrasound, followed by a validation test with E. coli O157:H7 at 60 °C. The cell morphology was observed with environmental scanning electron microscopy for samples treated at 40 °C and 60 °C. Physical quality attributes of the apple cider (pH, titratable acidity, °Brix, turbidity, and color) were compared for treated samples. The inactivation tests showed that sonication increased E. coli K12 cell destruction by 5.3‐log, 5.0‐log, and 0.1‐log cycles at 40 °C, 50 °C, and 60 °C, respectively. The additional destruction due to sonication was more pronounced at sublethal temperatures. At the lethal temperature of 60 °C, the rate of death by ultrasound was not significantly different compared with the thermal‐alone treatment. The inactivation of E. coli K12 with heat was described by 1st‐order kinetics, especially at 50 °C and 60 °C. For ultrasound treatments, concave upward survival curves were observed, which had a shape factor in the range of 0.547 to 0.720 for a Weibull distribution model. Extensive damage for ultrasound treated E. coli K12 cells, including cell perforation, was observed. Perforation is a unique phenomenon found on ultrasound‐treated cells that could be caused by liquid jets generated by cavitation. Titratable acidity, pH, and °Brix of the cider were not affected by ultrasound treatment. Minor changes in color and turbidity for ultrasound treated samples, especially for sonication at 40 °C for 17.7 min, were observed.
Ultrasonication and Fresh Produce (Cos lettuce) Preservation
Authors: Ajlouni Said , Sibrani Hatigoran , Premier Robert , Tomkins Bruce
Published: August 2010

Washing Cos lettuce in various sanitizers at different concentrations with and without ultrasonication (40 KHz) reduced the microbiological counts by 1 to 2.5 log colony‐forming units (CFU)/g immediately after washing. Ultrasonication of Cos lettuce in water, chlorinated water, peracetic acid, hydrogen peroxide, and their combinations at various temperatures (4 °C, 20 °C, 35 °C, 47 °C, and 50 °C) had no significant effects (P > 0.05) on the total or the psychrophilic counts during storage at 10 °C. The total count in Cos lettuce reached 9.74 ± 0.035 log CFU/g after ultrasonication (2 min at 50 °C) in chlorinated water (100 mg/L) and storage for 6 d at 10 °C. Extending the ultrasonication (40 kHz) of Cos lettuce for up to 20 min did not improve the bactericidal effect of ultrasonication. However, long‐time ultrasonication (20 min) caused significant (P < 0.05) damage to the quality of Cos lettuce tissues.
Keeping Quality of Desalted Cod Fillets in Consumer Packs
Authors: Magnússon Hannes , Sveinsdóttir Kolbrun , Lauzon Hélégne L. , Thorkelsdóttir ÁSa , Martinsdóttir Emilia
Published: August 2010

To meet modern consumer demands, it is important to be able to offer “ready‐to‐use” desalted cod in consumer packs with sufficient keeping quality at chill temperatures. The aim of this work was to investigate the effect of potassium sorbate, citric acid, and modified atmosphere packaging (MAP) in varying combinations on the keeping quality of desalted cod fillets in consumer packs. After desalting, cod fillets were packed in trays and stored at 1.0 ± 0.2 °C for up to 33 d. The fillets were packed in open bags, in modified atmosphere (MA) (CO2/N2/O2:75/ 20/5) only, or in MS following a potassium sorbate and/or a citric acid treatment. The rate of microbial growth was by far fastest in fillets in open bags. MAP alone decreased the growth rate considerably and still further decrease was obtained in MAP fillets treated with citric acid and/or sorbate solutions. The concurrent effect of these treatments was distinct. Quantitative descriptive analysis (QDA) was used to assess cooked samples. During the 1st d of storage, the samples were described by sweet and butter odor, salt taste, clammy and rubberlike texture, which became less evident with increasing storage time, but differed by groups and the least in samples treated with sorbate and citric acid/sorbate solutions. Sensory spoilage attributes and total volatile bases (TVB‐N)/trimethylamine (TMA) measurements correlated well with microbial counts. Use of MAP increased the shelf life from 6 to 10 d to 18 to 24 d, MAP and citric acid to 24 to 28 d, while the addition of sorbate to MAP fillets extended the shelf life to at least 33 d.
Hydroxypropyl‐Distarch Phosphate from Potato Starch Increases Fecal Output, but Does Not Reduce Zinc, Iron, Calcium, and Magnesium Absorption in Rats
Authors: Ebihara Kiyoshi , Nakai Yuka , Kishida Taro
Published: August 2010

The physiological effects of hydroxypropyl‐distarch phosphate (HDP) were examined in rats. Male rats were fed a fiber‐free purified diet containing gelatinized potato starch (PS, 50 g/kg diet) that was not modified chemically or gelatinized, chemically modified potato starch (CMS, 50 g/kg diet) for 21 d. PS was used as the control. Six kinds of gelatinized HDP from potato with 2 different degrees of hydroxypropylation and 3 different degrees of cross‐linking were used as CMS. The wet weight and moisture of the fecal output of the rats fed highly hydroxypropylated HDP was 70% and 30% greater, respectively, than that in the control rats. The weights of the cecal wall and content of the rats fed highly hydroxypropylated HDP were 15% and 30% higher, respectively, than those of the control rats. The apparent absorption of Mg was 15% higher in the rats fed highly hydroxypropylated HDP than in the control rats. On the other hand, the apparent absorptions of Ca, Zn, and Fe were not affected by the diet. Cross‐linking did not influence the above‐mentioned physiological effects of HDP. Fecal excretion of bile acids and the plasma cholesterol concentration were not affected by the diet. These results show that the physiological effects of HDP depend on the hydroxypropylation but not the cross‐linking.
Solution Enhancement and Post‐enhancement Storage Effects on the Quality, Sensory, and Retail Display Characteristics of Beef Triceps Brachii Muscles
Authors: Baublits R. Ty , Pohlman Fred W. , Brown A. Hayden , Yancey Ed J. , Johnson Zelpha B. , Dias‐Morse Palika
Published: August 2010

Beef triceps brachii muscles (6 d postmortem; n= 15; muscle sections n= 45) were sectioned into 3rds and allocated to 1 of 3 treatments. The treatments were untreated (CNT), or injected at a 12% pump rate with either tap water‐only (H2O) or a solution comprising tetrasodium pyrophosphate and sodium chloride (TSPP/ NaCl) at 0.4% and 1.0% target final product weight concentrations, respectively. Each muscle (comprising all 3 treatments) was then allocated to 2, 14, or 28 d of vacuum‐packaged 1°C storage. Purge losses during storage were greatest (P < 0.05) for H2O muscles and least (P < 0.05) for TSPP/NaCl muscles. Purge losses also increased (P < 0.05) from 2 d to 14 d of storage. Steaks enhanced with TSPP/NaCl had less (P < 0.05) free water and lower (P < 0.05) cooking losses than either CNT or H2O steaks. Storage duration did not affect (P > 0.05) Warner‐Bratzler shear force (WBS) or sensory tenderness, but juiciness decreased (P < 0.05) with increased storage duration. While storage duration did not impact (P > 0.05) instrumental color characteristics, aerobic plate counts generally increased during storage. The TSPP/NaCl steaks had lower (P < 0.05) WBS values and improved (P < 0.05) sensory tenderness and juiciness characteristics compared with CNT or H2O steaks. While CNT steaks had greater (P < 0.05) L* values (lightness) than TSPP/NaCl steaks, TSPP/NaCl steaks had similar (P > 0.05) oxymyoglobin proportions (630/580 nm) and a* values (redness) as CNT steaks. These results suggest enhancement with TSPP/NaCl can improve triceps brachii yield and palatability characteristics. Increased post‐enhancement storage did not impact or worsened palatability while increasing purge losses, suggesting general deleterious effects of increased postmortem storage for this muscle.
Increased Colocalization of Corticotropin‐Releasing Factor and Arginine Vasopressin in Paraventricular Neurones of the Hypothalamus in Lactating Rats: Evidence from Immunotargeted Lesions and Immunohistochemistry
Authors: Walker C.‐D. , Tilders F. J. H. , Burlet A.
Published: August 2010

In lactating female rats, tonically elevated glucocorticoid secretion is accompanied by blunted stress responsiveness, reduced expression of hypothalamic corticotropin‐releasing factor (CRF) mRNA and modest increases in arginine vasopressin (AVP) expression in the paraventricular nucleus (PVN). To determine the relative contribution of CRF and AVP to parvocellular function, we performed selective CRF (CRF‐Tx) or AVP (AVP‐Tx) lesions in the PVN neurones of ovariectomized virgin or lactating females (day 2 of lactation) by using ricin A associated with monoclonal antibodies directed towards CRF or AVP. We also performed double immunohistochemical labelling of CRF and AVP in the PVN of control rats injected with immunoglobulin (Ig)Gs associated with the ricin A (IgG‐Tx). Brains were collected 12 days after the lesion and processed forin situhybridization of CRF and AVP mRNA or for double fluorescence CRF and AVP immunohistochemistry. We found that lactating females exhibit a high degree of CRF and AVP colocalization in parvocellular PVN neurones, hypothalamic processes and median eminence terminals compared to virgins. While CRF mRNA is significantly reduced in lactating rats, AVP mRNA and protein levels are greatly enhanced in parvocellular PVN neurones during lactation. Hypothalamic CRF or AVP ricin‐A lesions significantly reduced both CRF and AVP expression (15–35% decrease) as well as peptide immunoreactivity in PVN neurones in both groups of females. The specificity of the lesions varied between virgins and lactators since in virgin females, AVP‐Tx did not affect CRF mRNA expression whereas in lactating females, this same lesion significantly reduced CRF mRNA expression, suggesting that parvocellular PVN neurones are more sensitive to the effects of the lesions during lactation. In both virgins and lactators, lesion with CRF‐Tx tended to increase AVP mRNA expression; however, in virgins, parvocellular PVN neurones were possibly compensating for the loss of CRF synthesis by increasing AVP expression and immunoreactivity. We conclude that lactation is associated with a high degree of CRF and AVP colocalization in parvoPVN neurones and that the increased AVP production in these neurones increases their sensitivity to immunotargeted lesions. The opposite regulation of CRF and AVP gene expression during lactation might provide a useful model to study differential sensitivity to glucocorticoid feedback or hypothalamic activation of transcription factors.
Efferent Neural Projections of Angiotensin Receptor (AT1) Expressing Neurones in the Hypothalamic Paraventricular Nucleus of the Rat
Authors: Oldfield B. J. , Davern P. J. , Giles M. E. , Allen A. M. , Badoer E. , McKinley M. J.
Published: August 2010

Angiotensin II acts within the hypothalamic paraventricular nucleus (PVN) to help mediate a number of autonomic and endocrine responses. Evidence is sparse in regard to the particular neuronal cell groups that exhibit angiotensin II type 1 receptors within the PVN, and does not exist in relation to specified efferent neuronal populations in the nucleus. In the present experiments, retrogradely transported neuronal tracers were utilized in conjunction with immunohistochemistry using a well characterized polyclonal antibody raised against a decapeptide sequence at the carboxy terminus of the AT1 receptor, to determine whether it is preferentially distributed amongst different efferent populations within the PVN. The AT1 receptor is not associated with neurones in the PVN that project axons to the spinal cord, dorsomedial or ventrolateral medulla but coexists strongly with neurones in the anterior parvocellular division of the nucleus which direct axons to the median eminence. Such neurones often contain corticotropin releasing factor. These findings highlight the role that angiotensin II and AT1 receptors in the PVN may play in the mediation of responses to stress.
Deviant Forms of Aggression in Glucocorticoid Hyporeactive Rats: A Model for ‘Pathological’ Aggression?
Authors: Haller J. , Van De Schraaf J. , Kruk M. R.
Published: August 2010

Deviant forms of aggressiveness have been associated with low plasma glucocorticoid concentrations in humans. Here, we report data on the development of aggressive behaviour in rats in which glucocorticoid secretion was inhibited by adrenalectomy. Such rats were compared with both sham operated rats and adrenalectomized rats in which the fight‐induced elevation of plasma glucocorticoids was mimicked by acute injections. Low and stable corticosterone plasma concentrations were maintained by subcutaneous glucocorticoid pellets in adrenalectomized rats. The development of aggressive behaviour was followed over three trials performed at 2‐day intervals. Adrenalectomy lead to high aggressiveness already at the first encounter, a decreased threatening (attack signalling) behaviour and a change in attack targeting. While control rats targeted biting attacks towards less vulnerable dorsal parts of the opponent's body, adrenalectomized rats attacked the head frequently. Corticosterone injections that mimicked the fight induced adrenocortical reaction abolished this behavioural pattern. Thus, a reduced responsiveness of the adrenocortical system may be causally linked to deviant forms of aggression in rats.
Restriction of Fetal Growth has a Differential Impact on Fetal Prolactin and Prolactin Receptor mRNA Expression
Authors: Phillips I. D. , Anthony R. V. , Simonetta G. , Owens J. A. , Robinson J. S. , McMillen I. C.
Published: August 2010

Prolactin is present in the fetal circulation and prolactin receptors are expressed in a wide range of fetal tissues. The factors which regulate the synthesis and secretion of prolactin, and the expression of its receptors before birth, are poorly understood. We have investigated whether experimental restriction of placental growth in the sheep has an impact on the prolactin axis in the growth restricted fetus. The majority of uterine endometrial caruncles were removed before pregnancy in 10 ewes (placental restriction; PR group). Placental, fetal liver and kidney weights were reduced in the PR compared to the control group (n = 10). The ratio of fetal prolactin mRNA : 18S rRNA was significantly lower (P < 0.01) in the PR group (1.83 ± 0.45, n = 6) than in the control group (4.11 ± 0.54, n = 6). The ratio of prolactin mRNA : 18S rRNA in the fetal pituitary was positively correlated with fetal and with placental weight. Using stepwise linear regression, it was determined that the level of fetal prolactin mRNA : 18S rRNA expression was best described (as judged by the maximum adjusted R2) by prolactin mRNA: 18 S rRNA = − 3.0378 + 0.17 PO2 + 2.772 glucose (adjusted R2 = 0.765, F = 17.53, P < 0.001). Fetal plasma prolactin concentrations were significantly reduced (P < 0.05) in the PR group compared to control animals between 109 and 141 days gestation. Fetal prolactin receptor (PRLR) mRNA transcripts encoding long (PRLR1) and short forms (PRLR2) of PRLR were present in the liver and kidney of animals in the PR and control groups at 140–141 days gestation. PR did not alter the levels of PRLR1 or PRLR2 mRNA in the fetal liver or kidney. The suppression of the synthesis and secretion of prolactin in the growth restricted fetus may limit the action of prolactin on the growth and metabolism of key fetal organs during suboptimal intrauterine conditions
Development of Luteinizing Hormone Releasing Hormone Neurones
Authors: Wray S.
Published: August 2010

This review concentrates on some of the recent discoveries and future questions relevant to the development of the neuroendocrine luteinizing hormone releasing hormone (LHRH) cells. Neuroendocrine LHRH cells originate outside the central nervous system, in the nasal placode, and thereafter migrate into the forebrain during prenatal development. It is this population of LHRH cells that is responsible for reproductive function, becoming integral members of the hypothalamo‐pituitary‐gonadal axis postnatally. Disruption of the development of this system results in reproductive dysfunction. Increasing our understanding of LHRH neuroendocrine cells establishes conditions where we can look with greater precision at the mechanisms controlling reproductive development, both activation and failure. In addition, the ability to manipulate the molecular and cellular biology of the LHRH system opens the route to understanding critical neurobiological issues such as phenotypic commitment, axonal path finding and mechanisms involved in neuronal migration. Each of the topics is discussed in turn and potential mechanisms controlling the development of the neuroendocrine LHRH system are indicated.
Effects of Testosterone on the Electrical Properties and Nicotinic Transmission of the Major Pelvic and Coeliac Ganglion Neurones
Authors: Félix B. , Catalin D. , Miolan J.‐P. , Niel J.‐P.
Published: August 2010

The effects of testosterone on the electrical properties and nicotinic activation of prevertebral ganglion neurones were investigated in vitro on the male rat major pelvic ganglion and rabbit coeliac ganglion. The electrical activity of the neurones was recorded using intracellular recording techniques. Nicotinic activation was triggered for neurones of the major pelvic ganglion by stimulating the hypogastric, pelvic and cavernous nerves and for coeliac neurones by stimulating the splanchnic nerves. Testosterone modified the resting membrane potential of neurones in the major pelvic ganglion by triggering a slow depolarization, and was without significant effect on the resting membrane potential of coeliac ganglion neurones. In neurones of the major pelvic and coeliac ganglia, testosterone had no significant effect on the firing pattern, on the characteristics of the action potential (firing threshold, duration, overshoot) and on the after‐hyperpolarization (amplitude and duration). Testosterone affected, in opposite ways, the nicotinic activation of neurones of the two prevertebral ganglia. In the major pelvic ganglion, testosterone triggered an increase in the amplitude of excitatory postsynaptic potentials induced by stimulation of the hypogastric, pelvic and cavernous nerves with a single pulse, revealing a facilitation of nicotinic activation. On coeliac ganglion neurones, testosterone elicited a decrease in the amplitude of excitatory postsynaptic potentials induced by stimulation of the splanchnic nerves, indicating an inhibition of nicotinic activation. Our study shows that testosterone acts differently on neurones of prevertebral ganglia involved in the nervous control of different functions, its facilitatory action being exerted on neurones of the major pelvic ganglion which is particularly involved in the control of the urogenital tract. Our study reinforces the concept, derived from neuroanatomical and pharmacological studies, of the major pelvic ganglion as a major peripheral target for testosterone.
Quantitative Evaluation of Synaptophysin‐Like Immunoreactive Nerve Terminals or Varicosities in Anterior Pituitary of Normal and Adrenalectomized Rats
Authors: Liu Y.‐Y. , Ju G.
Published: August 2010

Our previous studies have demonstrated the presence of substantial amounts of peptidergic nerve fibres in the anterior pituitary of monkeys, dogs and rats. The nerve fibres increase in number following adrenalectomy and ovariectomy. Furthermore, typical synapses between nerve fibres and anterior pituitary glandular cells have been demonstrated in the dog and rat, strongly suggesting the existence of some direct neural regulation. However, it is difficult to evaluate the functional importance of synapses or nerve terminals in general, unless quantitative data can be provided. The present study therefore aimed to quantify the number of terminals or varicosities in the anterior pituitary of the rat and its changes following adrenalectomy. Nerve terminals or varicosities were identified by their synaptophysin (SYN)‐like immunoreactivity. Under the light microscope, SYN‐like immunoreactivity appeared mainly as dots, mostly among glandular cells, and some were found distributed along blood vessels. SYN‐like immunoreactive dots were found under the electron microscope to be localized in the nerve terminals or varicosities. Synapse‐like contracts between the nerve terminals or varicosities and the glandular cells could be identified. The number of SYN‐like immunoreactive nerve terminals or varicosities was then counted under the light microscope. There were as many as 12424.2 ± 2941.5 and 20700.4 ± 6128 (mean ± SD) nerve terminals or varicosities per anterior pituitary of normal and adrenalectomized rats, respectively. The increase in number following adrenalectomy was statistically significant. It is concluded that nerve fibres may play an important role in the regulation of the anterior pituitary.
Pituitary Function and the Somatotrophic System in Patients with Idiopathic Parkinson’s Disease Under Chronic Dopaminergic Therapy
Authors: Schaefer S. , Vogt T. , Nowak T. , Kann P. H. ,   on behalf of the German KIMS board
Published: August 2010

Idiopathic Parkinson’s disease and dopaminergic medication may influence pituitary hormone secretion. The present study aimed to reveal any abnormalities of the somatotrophic system induced by the disease itself and/or the dopaminergic therapy. Investigations of other pituitary hormones under basal and stimulated conditions, as well as an analysis of body composition, were also performed. This was a controlled diagnostic study in which luteinising hormone‐releasing hormone, thyroid‐releasing hormone, corticotrophin‐releasing hormone and arginine hydrochloride were administered to ten patients with idiopathic Parkinson’s disease under dopaminergic medication. Basal and stimulated hormone concentrations and bioelectrical impedance analyses were compared with those of healthy, age‐matched controls. Basal growth hormone (GH) at −30 and 0 min was higher in Parkinsonian patients (2.74 ± 3.79 ng/ml versus 0.53 ± 0.10 ng/ml and 2.12 ± 2.44 ng/ml versus 0.51 ± 0.03 ng/ml; P < 0.05). The area under the GH curve after stimulation was greater in Parkinsonian patients (502.4 ± 202.6 ng × min/ml versus 312.0 ± 98.5 ng × min/ml; P < 0.05), depending on higher basal GH levels, rather than a greater arginine response. No differences in insulin growth factor (IGF)‐1 or IGF‐BP3 concentrations were detected. There were no differences between the groups in basal and stimulated gonadotrophic, corticotrophic and thyrotrophic function, or body composition. Prolactin was below the detection limit in the patients during the course of the study. Parkinsonian patients experience marked hypoprolactinaemia and repeated stimulation of GH secretion during chronic dopaminergic therapy. Our findings suggest a peripheral GH resistance in these chronically‐treated patients.
A PREPARATIVE METHOD FOR THE ISOLATION OF CARBOXYL TERMINAL TRYPTIC PEPTIDES FROM PROTEINS
Authors: Hargrave Paul A. , Wold Finn
Published: August 2010

A method has been developed for isolating glycinamide‐blocked carboxyl‐terminal tryptic peptides from proteins. It has been successfully applied to four proteins of known carboxyl‐terminal sequence, yielding the carboxyl‐terminal peptide in up to 68% yield. The separation method is designed to be selective for the desired peptide and to handle any quantity of protein.
STUDIES ON MOLD DEXTRANASES: Some Physicochemical Properties and Substrate Specificity of Dextranases Obtained from Aspergillus carneus and Penicillium luteum
Authors: Hiraoka Nobutsugu , Tsuji Hikoji , Fukumoto Juichiro , Yamamoto Takehiko , Tsuru Daisuke
Published: August 2010

Some physicochemical properties and substrate specificity of the highly purified preparations of dextranases from Aspergillus carneus and Penicillium luteum were investigated. The values of 71,000 and 46,000, respectively, were obtained for the molecular weight of dextranases of Asp. carneus and Pen. luteum by ultracentrifugal analysis, and their sedimentation coefficients, S20,w, were 4.8 S and 4.1 S, respectively. The amino acid composition of these two enzymes was very similar except for the contents of histidine, arginine, threonine and serine. Comparison of enzymatic digestion products from isomaltodextrins and their reduced derivatives (sugar alcohols) indicated that Asp. carneus dextranase attacked the substrates, liberating isomaltotriose and, to a lesser extent, isomaltose from their non‐reducing ends. It was concluded that Pen. luteum dextranase split the substrates primarily by removing an isomaltosyl unit and, to a somewhat lesser extent, an isomaltotriosyl unit from their non‐reducing ends.
A NEW METHOD OF CALCULATING EVOLUTIONARY RATES OF PROTEINS APPLIED TO INSULIN AND C‐PEPTIDES
Authors: Markussen Jan , Vølund Aage
Published: August 2010

The optimal number of differences between sequences for phylogenetic studies is derived, and the developed theory is used to show that a sequenced peptide assumed to be duck proinsulin C‐peptide had the expected differences from the mammalian C‐peptides. The theory is used to separate the guinea pig insulin from other insulins, indicating that the pressure of selection on guinea pig insulin has been significantly lower than that on other insulins. Finally, calculation of the half‐life of a protein as a measure of the pressure of selection rather than the evolutionary rate constant is proposed.
STUDIES ON POLYPEPTIDES
Authors: Romovacek Hana , Drabarek Stefania , Kawasaki Koichi , Dowd Susan R. , Obermeier Rainer , Hofmann Klaus
Published: August 2010

The synthesis by fragment condensation of a nonatricontapeptide corresponding to positions 66–104 of the proposed amino acid sequence of the enzyme ribonuclease T1 is described. It is shown that complex peptides corresponding to the C‐terminal region of the T1 enzyme are soluble in 8 M pH 10.7 urea and that purification can be achieved by gel filtration in this solvent
IMMUNOCHEMICAL INVESTIGATION ON HUMAN CERULOPLASMIN. PARTIAL EXPLANATION OF THE «HETEROGENEITY»
Authors: Løwenstein Henning
Published: August 2010

Human ceruloplasmin from fresh serum has been purified by chromatography on hydroxyapatite and Con A‐Sepharose. Quantitative immunoelectrophoretic analysis of fresh serum, stored serum and fractions from the different purification steps for human ceruloplasmin has been carried out. A combination of the latter, advanced technique with amino acid analysis, molecular weight determination by size chromatography, urea treatment, staining for oxidase activity and enzymatic proteolysis, has revealed that: 1) human ceruloplasmin is a heterogeneous mixture of two glycoproteins (x) differing only in their carbohydrate content and 2) the protein part contains at least one very labile peptide bond which upon enzymatic hydrolysis gives rise to peptides with molecular weights of 93,000 (y) and 24,000 (z) dalton, respectively. The two glycoproteins are immunochemically identical. The y peptide is immunochemically partially identical, and the z peptide immunochemically non‐identical, with the parent molecule. The y and z peptides are non‐identical. On the basis of these observations a simplified two‐dimensional model of human ceruloplasmin is proposed.
ON THE REACTIVITY OF CARBOXYL GROUPS OF RIBONUCLEASE‐A IN ANHYDROUS METHANOL
Authors: Acharya A. Seetharama , Vithayathil Paul J.
Published: August 2010

The esterification of Ribonuclease‐A in methanol/0.1 M hydrochloric acid has been studied by measuring the decrease in the number of titratable groups of the protein and estimating the amount of methanol incorporated. Esterification of nearly five of the 11 free carboxyl groups of the protein resulted in almost complete inactivation of the enzyme. The initial products of esterification have been chromatographed on Amberlite columns, and five partially active methyl ester derivatives of Ribonuclease‐A have been isolated. The dimethyl ester, the initial product of esterification with reduced catalytic activity, has the carboxyl groups of Glu‐49 and Asp‐53 modified. Even in the non‐aqueous solvent, as in the native structure of the protein in aqueous solution, these carboxyl groups are the fast reacting ones. Subsequently, the esterification reaction appears to proceed preferentially at the C‐terminal region of the molecule. Comparison of the reactivities of carboxyl groups of Ribonuclease‐A in acidic methanol to that known in aqueous solutions (with carbodiimides) suggests that the structure of Ribonuclease‐A in the non‐aqueous solvent resembles, at least in part, the structure in aqueous environment.
ELECTRON PARAMAGNETIC RESONANCE STUDIES ON SPIN‐LABELLING OF PEPSIN: EFFECTS OF TEMPERATURE, pH AND UREA ON ITS CONFORMATION
Authors: Aoshima Hitoshi , Naito Akira , Hatano Hiroyuki
Published: August 2010

Pepsin was spin‐labelled with N‐(1‐oxyl‐2,2,6,6‐tetramethyl‐4‐piperidyl)bromo‐acetamide, possibly at the active site, at a β‐carboxyl group of a reactive aspartic acid. The spectrum of the spin‐labelled pepsin showed that the spin probe was strongly immobilized (correlation time ≥10‐a sec). Spin‐labelled pepsin was thermally denatured at various temperatures and electron paramagnetic resonance (e.p.r.) spectra were taken at various times. Rates of denaturation estimated from the e.p.r. spectra at various temperatures showed that the enthalpy and entropy of thermal denaturation of spin‐labelled pepsin at pH3.5 were 48.0 ± 4.9 kcal/mole and 214.7 ± 14.5 e.u. respectively. Addition of conc. NaOH or 1 M acetate buffer at pH 6.0 sharpened e.p.r. spectra of the spin‐labelled pepsin, indicating that the spin probe became mobilized by alkaline denaturation. Addition of urea caused unfolding of the protein which increased with the urea concentration, although only slight transition of conformational changes was observed in the e.p.r. spectra.
STUDIES ON PITUITARY LACTOGENIC HORMONE. THE PRIMARY STRUCTURE OF THE PORCINE HORMONE*
Authors: Li Choh Hao
Published: August 2010

The complete amino acid sequence of porcine lactogenic hormone has been elucidated. It consists of 199 amino acid residues with three disulfide bridges formed by residues 4–11, 58–174, and 191–199. The two tryptophan residues are in positions 91 and 150. A high degree of homology exists between the known structure of ovine prolactin and the porcine lactogenic hormone. This led to a re‐examination of residues 82–90 in the ovine prolactin structure, where an extra leucine was found in position 88.
STUDIES ON THE PRIMARY STRUCTURE OF BISON PANCREATIC RIBONUCLEASE
Authors: Muskiet Frits A. J. , Welling Gjalt W. , Beintema Jaap J.
Published: August 2010

Bison pancreatic ribonuclease was isolated by affinity chromatography. Thermolysin and tryptic digestion of denaturated protein, and subtilisin digestion of native protein yielded peptides, which were purified and submitted to amino acid analysis.

These peptides, together with partial sequence data obtained by Stewart & Stevenson (16) overlap the entire amino acid sequence of bison ribonuclease. No differences with bovine ribonuclease were found, although there may be differences in state of amidation of some residues.
THE HAEM‐ACCESSIBILITY IN LEGHAEMOGLOBIN OF LUPINUS LUTEUS AS OBSERVED BY PROTON MAGNETIC RELAXATION
Authors: Vuk‐Pavlović S. , Benko B. , Maričic S. , Lahajnar G. , Kuranova I. P. , Vainshtein B. K.
Published: August 2010

Using the solvent‐protons' longitudinal magnetic relaxation rates (p.m.r.) for Lupinus luteus leghaemoglobin derivatives the accessibility of the haem has been evaluated by our “stereo‐chemical p.m.r. titration” method with nonexchangeable protons of aliphatic lower alcohols in otherwise deuterated solutions. The haem in leghaemoglobin is more accessible and its protein environment more flexible compared with vertebrate haemoglobins. The correlation time in aquometleghaemoglobin aqueous solution has been determined by measuring the frequency dispersion of the p.m.r. rates between 6.1 and 93 MHz. Taking into account the measured value of τc = (7.7 ± 0.5) × 10‐10 s the iron‐to‐proton inter‐spin distances have been calculated. The significance of these distances as well as the electronic g‐factor anisotropy for elucidation of fine structural details of the haem‐environment are discussed.
THERMAL FRAGMENTATION OF ESCHERICHIA COLIβ‐GALACTOSIDASE.
Authors: Marinkovic Dobrivoje V. , Marinkovic Jelka N. , Tang Jordan
Published: August 2010

Carboxymethylated Escherichia coli β‐galactosidase EC 3.2.1.23 could be broken to polypeptides of fairly uniform size (average molecular weight about 22,000 daltons) by heating for ≤8 h at 100°C and pH 7.5 in 8 M‐urea. Using phosphocellulose chromatography in NaCl‐urea gradients, the resulting polypeptide mixture could be resolved in three fractions essentially homogeneous by disc gel electrophoresis in urea at several pH values, and by isoelectric focusing. One of these fractíons was active as α‐donor in in vitro complementation of β‐galactosidase activity with Escherichia coli mutant M15; this activity was largely retained after CNBr cleavage. All three fractions carried arginine as carboxyl‐terminal amino acid. No significant amount of any specific amino could be detected in NH2‐terminal position.
INVESTIGATION ON THE REACTION OF MOLYBDENUM WITH COLLAGEN IN VIVO
Authors: Bibr BedŘIch , Deyl ZdeněK , Lener Jaroslav , Adam Milan
Published: August 2010

Male Wistar rats were treated with Mo in a single dose or over a period of 84 days. The course of molybdenum excretion after a single dose was expressed mathematically. It was shown that various tissues produced different excretion curves and a high retention was observed only in skin. In long‐term treatment it was possible to ascribe this retention to direct binding of molybdenum to collagen. In contrast to some other heavy metals molybdenum causes a decrease of collagen stability. The mechanism of this effect is discussed.
SHORT‐AND LONG‐RANGE SPECTRAL PERTURBANTS: DEPENDENCE ON SOLVENT CONCENTRATION AND CHROMOPHORE STRUCTURE
Authors: Bello J. , Bello H. R.
Published: August 2010

Some perturbing solvents appear to be short‐range perturbants at 20% concentration, but long‐range at 40%, based on Δε/ε for phenols and their ethers. The apparent long‐ or short‐range character of a solvent also depends on the chromophore. It is possible that perturbants, which are apparently of long‐range character at 20% concentration, are really of short‐range character because they interact rather similarly with phenols and their ethers. For ethers Δε/ε increases faster than for phenols with increasing concentrations of some perturbants.
ARGININE MODIFICATION IN KUNITZ BOVINE TRYPSIN INHIBITOR THROUGH 1, 2‐CYCLOHEXANEDIONE*
Authors: Menegatti Enea , Ferroni Roberto , Benassi Carlo A. , Rocchi Raniero
Published: August 2010

Arginine residues (5.5 out of 6) of the trypsin‐kallikrein inhibitor from bovine organs (Kunitz inhibitor) were selectively modified by reaction with 1, 2‐cyclo‐hexanedione in sodium borate buffer, pH 9.0. The modified inhibitor is still highly active in inhibiting trypsin and chymotrypsin at 1:1 inhibitor: enzyme molar ratio and full inhibition was achieved at slightly higher molar ratio. The extent of correct refolding, upon reoxidation, of the reduced, arginine‐modified inhibitor is diminished and regeneration of two arginines occurred under the reduction conditions.

The stability constants and the standard‐free energies of binding of the complexes between trypsin, or chymotrypsin, and the native, the arginine‐modified and the reduced and reoxidized arginine‐modified inhibitor have been determined from inhibitory assays.
DOPAMINE β‐MONOOXYGENASE
Authors: Skotland Tore , Ljones Torbjørn
Published: August 2010

A simple and sensitive method for determination of amide groups in proteins is presented. The method employs a combination of a simple microdiffusion technique and the indophenol method for ammonia determination. A similar ammonia analysis was used for total nitrogen determination after Kjeldahl digestion. The results from the amide group and total nitrogen analyses, together with previously published amino acid and carbohydrate analyses, were used to estimate a value of 12.4 for the specific extinction coefficient, E1%280 nm, of dopamine β‐monooxygenase from bovine adrenal medulla.
THE ACYLATION OF BOVINE SERUM ALBUMIN WITH DIACETYLCYCLOSERINE
Authors: Howard John Charles
Published: August 2010

The reaction of the amino groups of bovine serum albumin (BSA) with diacetyl‐cycloserine (I) at pH 7.2–9.0 proceeded with both acylation by the diacetyl‐β‐aminooxy‐D‐alanyl (DAA) group and acetylation. The number of DAA groups was determined by their conversion to cycloserine (III) which can be accurately measured in micromolar amounts. The method was developed using the model compound diacetyl‐β‐aminooxy‐D‐alanine butyl amide (II) which was converted to β‐aminooxy‐D‐alanine butyl amide (IV) by methanolysis and then to cycloserine by basic ring‐closure of IV. Calculations using results obtained by this method combined with the experimentally determined number of free amino groups in the modified BSA indicated that the reaction of excess I with BSA effected the acetylation of about 35 and the acylation with DAA groups of about 22 of the 59 amino groups. These findings were supported by experiments demonstrating that the amount of acetic acid formed by hydrolysis of the modified BSA was approximately that predicted from the results of the cycloserine analyses.
PURIFICATION OF MILK‐CLOTTING PROTEASE FROM BACILLUS MESENTERICUS STRAIN 76
Authors: Mesrob B.K. , Stoeva St.P. , Vassileva R.A.
Published: August 2010

A two‐step procedure for the isolation of pure enzyme from Bacillus mesentericus strain 76 is developed. Ion‐exchange chromatography on CM‐Sephadex‐C50 is used as a second step after precipitation from ethanol. The pure enzyme preparation, obtained after gel filtration on Sephadex‐G25 of the ion‐exchange chromatographically separated product, possesses the highest specific activity achieved. The homogeneity of the final preparation is proved by determination of the N‐terminal amino acid (Arg) and by SDS‐PAGDE (single disc).
A SURVEY OF ATOM PACKING IN GLOBULAR PROTEINS
Authors: Crippen Gordon M. , Kuntz I. D.
Published: August 2010

This survey of the atom packing in the high resolution X‐ray crystal structures of 21 proteins indicates that the atom density around a given central atom is determined primarily by its covalently bonded neighbors and proximity to the surface of the protein. Long‐range hydrophobic, hydrogen bonding, and electrostatic interactions are strictly of secondary importance. Both radial and angular atom densities were calculated about various central atoms of several residue types, averaged over all occurrences of the chosen residue type in all 21 proteins. Polar interactions, such as hydrogen bonding, involve on the average shorter distances than hydrophobic interactions do, and are more directional. Spatial segregation of polar and non‐polar atoms is never complete in long‐range interactions since the types of atoms are linked together covalently.
DISSOCIATION, AGGREGATION OF SESAME α‐GLOBULIN IN NONIONIC DETERGENT SOLUTION
Authors: Lakshm T. S. , Nandi P. K.
Published: August 2010

Nonionic detergents Triton X‐100 and Brij 36T induce dissociation and aggregation of the protein sesame α‐globulin above the critical micelle concentrations (cmc) of the detergents. Spectrophotometric titration in Triton shows no change in the pKInt value of the tyrosyl groups at 1 × 10‐3M detergent where both dissociation and aggregation of the protein are observed. Fluorescence measurement does not indicate any change in the environment of the tryptophan groups of the protein in Brij. Viscosity measurements show no major conformational change of the protein in the detergent solution. Binding measurements suggest that perhaps micelles of the detergent predominantly bind to the protein. The detergent micelles preferentially bind to the exposed hydrophobic surfaces of the protein subunits. The association of the protein detergent complex through electrostatic interaction is probably responsible for the formation of the aggregates.
STUDIES ON PITUITARY FOLLITROPIN
Authors: SAIRAM M.R. , LP CHOH HAO
Published: August 2010

The α and β subunits of human follitropin were isolated in a high state of purity. The tryptophan fluorescence of the native hormone and the isolated β subunit are different. The N‐terminus of the α and β subunits was identified as valine and aspartic acid respectively. While recombination of the isolated α and β subunits restores the electrophoretic mobility of the intact hormone, its receptor binding activity cannot be fully regenerated. Substitution of the human follitropin α by an ovine lutropin α subunit, to form a recombinant with the follitropin β subunit, generates a complex with 2–3 receptor binding activity of the native human follitropin and the same activity as ovine follitropin. Acylation of the intact hormone does not disrupt the quaternary structure but leads to complete inactivation. Acylation studies with the subunits suggests the crucial role of the ∍‐amino groups of the α subunit in determining biological activity.
TOBACCO LEAF MESOPHYLL PEROXIDASE Isolation, Purification and Some Properties
Authors: YUNG KUNG‐HING , WONG YUM‐SHING , CHOY YUEN‐MIN
Published: August 2010

Two isoperoxidases, P1 and P2, were isolated from tobacco mesophyll leaves by the combined methods of ammonium sulphate precipitation, PAGE and DEAE‐cellulose column chromatography. The pH optima for P1 and P2 were 7.0 and 7.5 respectively. The values of Km for H2O2 were 1.82 and 0.87 mM for P1 and P2. Results from this investigation suggest that these isoperoxidases differ in their electrophoretic, catalytic and physical properties.
CHEMICAL, BIOLOGICAL AND IMMUNOLOGICAL CHARACTERIZATION OF SEPARATED ELECTROPHORETIC COMPONENTS OF HUMAN CHORIONIC SOMATOMAMMOTROPIN (HCS)
Authors: TARLI P. , BOTTI R. , COCOLA F. , FABRIZI P. , NERI P.
Published: August 2010

Purified HCS preparations often show heterogeneity when assayed by electrophoresis, although they may be considered homogeneous according to other criteria (N‐terminal amino acid analysis, gel filtration and ultracentrifugation patterns, etc.). Using DEAE‐cellulose chromatography, the two main components corresponding to various degrees of hormone amidation were separated. The deamidation kinetic of the electrophoretically slower migrating component was studied and found to be sensitive to temperature and, to a lesser extent, to pH. Denaturing agents strongly increased the deamidation rate, while no effect was found by raising the ionic strength. No difference was evident between the two components and the parent hormone when compared (on a weight basis) in a specific HCS radioimmunoassay system. Even the precipitin reaction in agar gel with an anti‐human growth hormone (HGH) serum did not show variations in cross‐reactivity versus HGH. Moreover, groups of guinea pigs immunized with the single HCS components, gave antibodies reacting identically with the components and the parent HCS. The prolactin‐like activity of the two components differed when determined by the pigeon crop‐sac assay: the slower migrating component proved to be less active than the more deamidated one. On the other hand, the activity in two different radioreceptor assays (employing membranes from rabbit mammary gland and rat ventral prostate) was greater for the faster migrating component.
THE PREFERRED CONFORMATIONS OF PROTECTED HOMODI‐TO HOMOHEPTAMETHIONINE PEPTIDES A 1H N.M.R. Study in Deuterochloroform Medium
Authors: RIBEIRO ANTHONY A. , GOODMAN MURRAY , NAIDER FRED
Published: August 2010

Detailed analyses of the conformations of the homo‐oligopeptide series, Boc‐(L‐Met)n‐OMe n = 2–7, in deuterochloroform have been carried out with proton n.m.r. and IR spectroscopy. Well‐resolved high field n.m.r. spectra with assignments for the NH and α‐CH resonances of these homo‐methionine peptides are presented. Extensive n.m.r. concentration‐dependent chemical shift studies are combined with IR results to delineate the involvement of the various methionine NH protons in intra‐ and/or intermolecular hydrogen bonding. N.m.r. chemical shift dependencies with temperature and solvent, DMSO‐d6, are used to explore the strength of the hydrogen bonds for the various oligopeptides. At low concentrations, where peptide aggregation is absent, the dipeptide is found to be disordered. The tetra‐ to heptapeptides possess intramolecular hydrogen bonded seven‐membered rings at internal residues. The number of internal rings and the oligopeptide self‐association increase with increasing peptide chainlength. At intermediate concentrations associations of peptide molecules with folded structures occur with initial association at the C‐terminal region. At high concentrations, “in‐register” associated extended β structures are formed.
X‐RAY STUDY ON HOMO‐OLIGOPEPTIDES; t‐BOC(L‐NVA)6OME AND HCI‐ H(L‐NVA)6 OME
Authors: SPADON P. , DEL PRA A.
Published: August 2010

Observations of extended peptide chains, whose direction is perpendicular to the fiber axis (cross‐ßt‐structure) have hitherto been confined to fibrous proteins and to some synthetic polydisperse polypeptides of rather low molecular weight. This structure has now been found in some monodisperse linear homooligopeptides with aliphatic hydrocarbon side chains. X‐ray fiber diagrams of t‐Boc(L‐Nva)6OMe and HCl‐H(L‐Nva)6OMe show the characteristic reflections of this form. In addition, the good orientation of suitably prepared specimens has enabled a fairly complete determination of the hexapeptides unit cell to be made. Both molecules are packed in a pseudomonoclinic lattice whose dimensions are: a = 4.80, b = 59.0, c = 14.86 Å, (ß= 85 and a = 4.80, b = 58.50, c = 14.84 and ß= 86 for t‐Boc(L‐Nva)6‐OMe and HCl‐H(L‐Nva)6OMe respectively. It has not been possible from the available experimental data to establish whether the arrangement of the chains within the sheets is parallel or antiparallel.
HYDROXY‐ AND AMINO‐PROTECTION BASED ON THE 4‐DIMETHYLCARBAMOYLBENZYL GROUP
Authors: CHAUHAN VIR S. , RATCLIFFE STEVEN J. , YOUNG GEOFFREY T.
Published: August 2010

O ‐ 4 ‐ Dimethylcarbamoylbenzyl ‐ L ‐ tyrosine, N (e)‐4‐dimethylcarbamoylbenzyl‐oxycarbonyl‐L‐lysine, and simple derivatives, have been prepared. The protecting groups are markedly more stable to trifluoroacetic acid than are the unsubstituted benzyl analogues. N‐t‐Butoxycarbonyl‐O‐4‐dimethylcarbamoylbenzyl‐L‐serine (cyclohexylammonium salt) is also described.
ALTERNATIVE SYNTHESIS OF CORTICOTROPIN‐LIKE INTERMEDIATE LOBE PEPTIDE (HUMAN CLIP)
Authors: TURÁN ANDRÁS , BAJUSZ SÁNDOR
Published: August 2010

A novel synthesis of human CLIP, a peptide corresponding to the sequence of human ACTH‐(18–39) is described. The dodecapeptide chain was assembled by a combination of fragment condensation and stepwise synthesis while using Nα‐benzyloxycarbonyl and side chain tert.‐butyl‐derived protective groups combination. The final deprotection was performed by acidolysis in trifluoroacetic acid. The end product was purified by ion exchange chromatography on carboxy‐methyl cellulose.
CIRCULAR DICHROISM SPECTRA OF CLEAVAGE FRAGMENTS OF SOYBEAN TRYPSIN‐CHYMOTRYPSIN INHIBITOR
Authors: BIRK YEHUDITH , JIBSON MICHAEL D. , BEWLEY THOMAS A.
Published: August 2010

Circular dichroism spectra of biologically active fragments of Bowman‐Birk soybean inhibitor have been determined in acidic, neutral, and alkaline conditions. Neither fragment showed evidence of α‐helix or β‐structure. Negative dichroism above 260 nm has been assigned in both fragments primarily to disulfide bonds, with a minor contribution from tyrosine in a hydrophilic environment. The individual spectra of these fragments, and their sum between 230 and 340 nm have been compared with the spectra of the intact inhibitor and several structurally related proteins. Possible interactions which may give rise to CD bands in this region are discussed.