Direct determination of free methionine in soy-based infant formula

A method was developed for the direct determination of free methionine in soy-based infant formula, with analyte separation and quantitation by reversed-phase liquid chromatography (LC), and UV absorbance at 214 nm, respectively. Sample preparation required only dilution with mobile phase and syringe filtration. Using a 0.02M KH2PO4 mobile phase (pH adjusted to 2.9 with 85% o-phosphoric acid) and 0.7 mL/min flow rate, methionine eluted at approximately 8 min, and total run time was 14 min after column regeneration with acetonitrile-water. System linearity was demonstrated as peak area versus analyte concentration, ranging from 80 to 120% of the formula specification for free methionine (r > 0.999, and all residuals < 0.45%). Intermediate precision relative standard deviation values were < 1.5% for ready-to-feed and reconstituted powder samples, and recoveries ranged from 98.0 to 103.5% for inter-method comparison with an amino acid analyzer method. The limit of quantitation was 3 mg methionine/L in the "as fed" infant formula. Despite the relatively weak UV absorptivity of methionine, the 214 nm signal was sufficiently intense in the 30-65 mg/L (201-436 microM) range to afford quantitation by peak area proportionation versus a 2-point external standard calibration. This direct UV detection after reversed-phase LC separation provides a simple and accurate method for determining free methionine without derivatization.     

ETUDE DE LA FRAGMENTATION DE N-ARYL ET DE N-ALKYL AMINOCETONES EN SPECTROMETRIE DE MASSE

A Comparison of the chemical ionisation and elctron-impact mass spectra of alkyl and aryl aminoketones has bee carried out. This has revealed the particular stabilization of the N-aryl grops in all cases.     

Zinc solid-state NMR spectroscopy of human carbonic anhydrase: implications for the enzymatic mechanism

The pH dependence of the (67)Zn solid-state nuclear magnetic resonance spectroscopy of human carbonic anhydrase (CAII) has been investigated to characterize the nature of the fourth ligand. CAII, through the Zn(2+)-bound hydroxide, catalyzes the deceptively simple reaction: CO(2) + H(2)O <==> HCO(3)(-) + H(+). The accepted mechanism for CAII would predict that water would be bound to the Zn(2+) at pH 5 and hydroxide would be bound at pH 8.5. The measured values for the electric field gradient (EFG) or quadrupole coupling constant (Cq) for CAII are independent of pH within the limits of the experimental error, i.e., 9.8 +/- 0.2 MHz. The EFG interaction has been predicted by ab initio electronic structure calculations for water and hydroxide bound to the zinc, including various levels of hydrogen bonding. After comparing the predicted Cq's with the experimental values, we conclude that the species present from pH 5-8.5 is the hydroxide form. The NMR data presented here is not consistent with the accepted mechanism for CAII. We show that the NMR data is consistent with an alternative mechanism of CAII.     

Substituent effects in chemical carcinogenesis: Methyl derivatives of the benzacridines

An explanation for the different carcinogenic potencies observed among methyl derivatives of the angular benzacridines is given in terms of the tendencies of these compounds to undergo specific metabolic activating reactions analogous to those of polycyclic aromatic hydrocarbons. Theoretical reactivity indices representing these reactions correlate with the carcinogenic activities of these compounds.     

Adsorption kinetics and electrode processes

Prof. H. A. Laitinen (Urbana):We have used double layer capacity measurements to measure adsorption as a function of time during the life of slowly forming mercury drops. For many substances that are not extremely surface active, so that the solution concentration is 1 to 10 mM, the double layer capacity per unit area is essentially constant during the time interval of 5 to 15 sec of drop life. It appears, therefore, that adsorption equilibrium has essentially been attained during the later portions of drop life.Prof. P. Delahay:We have also used a similar method of following the differential capacity during drop life. I would agree that one can achieve conditions for which adsorption equilibrium with respect to bulk concentration is practically reached (high enough concentration and long drop time). But this is quite often not the case (drop time of 3–4 sec in polarography).Mr. G.C. Whitnack (China Lake, U.S.A.);In determination of nitrate esters in presence of phthalate esters we have observed considerable dropping of current height of nitrate ester wave before start of phthalate ester wave. Is this due to an adsorption process occurring at the DME?Prof. P. Delahay:It might well be, but I would hesitate to comment any further on this point because more information would be required. You probably have quite a complicated situation. Perhaps we can discuss the matter later?Prof. Dr. H. Fischer (Ettingen):Ich möchte hinzufügen, class es Fälle geben kann, in denen mcht allem der Blockierungseffekt don Grenzstrom verringert, sondern eine Veränderung des Diffusionsfilmes durch Sekundärreaktion des Inhibitors. Dies ist z.B. bei der Abscheidung von Wasserstoff an einer festen Elektrode (Fe) der Fall, wenn der Inhibitor sich spaltet in eine schwerlösliche Verbindung und ein Proton (RNH⁺ → [RN] + H⁺). Dies beobachtet man z.B, bei den Kationen von heterozyklischen Aminen (Acridin). Offentlich bildet sich eine diffusionshindernde Barriere aus.Prof. P. Delahay:The case of the hydrogen electrode is complicated, and I shall attempt to answer your question only in the case of an ideally smooth electrode. Then, diffusion toward the electrode (partially covered with an adsorbed substance) is little affected by adsorption because the size of “the holes” in the film is very small in comparison with the diffusion layer thickness. Of course, there is no diffusion where there is complete blocking.Prof. B. Breyer (Sydney):The importance of the chemical nature of the film adsorbed at the interface, which has been mentioned by Prof. delahay, seems to me to play a major part in the type of processes discussed. Thus it must be kept in mind that complex formation between the diachargeable ion and the adsorbed film might occur (cf. e.g. heyrovsky? and matyas, 1941). The fact that T1+ ion is little influenced by the presence of an adsorbed film at the electrode solution interface could then be partly explained by the notoriously low co-ordination tendency of that ion.Prof. P. Delahay:The difficulty involved in the “blocking” of the limiting current for thallium is due, I think, to the small size of this ion (large diffusion coefficient of Tl+ in comparison with other ions). Of course, complex ions can be relatively very bulky and this enhances “blocking”.Prof. E. Lange (Erlangen):I agree with prof. delahay that it is very important to investigate the connections between the adsorption and electrode reaction.This is easy for a steady state, e.g., each heterogeneous reaction between two phases that is accompanied by a transfer of ions or electrons, i.e. of electrical charges, from one phase to the other. In such a case, the Galvani tension does not change and the transfer between the two phases must be compensated by a corresponding transfer of charges within the phases.But in the non-steady state, also, an adsorption process may behave as an “electrode reaction” for instance, even an adsorption of a dipole molecule may cause a “transfer of charge” accompanied by a corresponding change of the Galvani tension. In this manner, it seems to me that for the non-steady state it is necessary to define precisely what one means by “electrode reaction”.Prof. P. Delahay:I entirely agree with Prof. i.ange about the necessity of clear definitions. I think that the fact that a steady state with respect to diffusion of the reductible or oxidizable substance has not been reached is not too serious because this scarcely affects the Galvani potential (large excess of supporting electrolyte).Variations in the amount of adsorbed organic substances indeed affect the Galvani potential (dipole orientation), but this effect is included in the dependence of the rate constant ks (at the standard potential) on the electrode coverage.Prof. N. Tanaka (Sendai, Japan).I am very grateful to Prof. delahay for his beautiful work on adsorption kinetics. I should like to make one comment in connection with the rotated dropping mercury electrode. The relation between log i and log t on the current — time curve was found to be 0.5 only in the absence of the surface-active substance. In the presence of surface-active substance, the slope of log i vs. log t changed at a certain point of the current — time curve. This can be explained when the slow adsorption of the surface-active substance on the surface of the electrode is taken into consideration.Prof. P. Delahay:Even for stirred solutions, adsorption equilibrium is not reached very rapidly. A simple calculation based on a model of the Nernst diffusion layer shows that perhaps 1–5 sec are required. Your conclusion is, therefore, quite correct.Prof. W. Kemula (Warsaw):We have recently published that, in several cases, the addition of extremely small concentrations of organic surface-active substances provokes at first a rise of the diffusion current, this current then being suppressed by additional quantities of the substance.     

Semisimilarity of molecular bodies: Scaling–nesting similarity measures

A simple, new technique for the evaluation of the similarity of molecular shapes is presented. The concept of semisimilarity (asymmetric similarity) is applied within the topological–geometrical framework of scaling–nesting similarity measures of molecules. The practical application of these similarity measures is illustrated by the examples of the three-dimensional formal bodies of electronic charge densities of a set of simple molecules. © 1994 John Wiley & Sons, Inc.     

Contribution to the study of the mechanism of curing of unsaturated polyester resins

The mechanism of copolymerization of monomethyl and dimethyl maleates and fumarates with styrene was studied by analysis of the conformation of the acid units of the resulting copolymers. The absorption bands for C?O stretching and OH stretching in the spectra of the copolymers are fully identical. They are quite different from the spectra of the copolymers obtained from maleic anhydride and styrene that are subsequently treated with absolute methanol to give the monoester which is then esterified with diazomethane to give the diester. The acid units of the copolymers derived from maleic anhydride exist in a gauche configuration; copolymers derived from fumaric units exist in a trans conformation. The identity of copolymers derived from maleic units with those derived from fumaric units but not with those derived from maleic anhydride indicates that the first step in the copolymerization of the maleic units is an isomerization to fumaric units, which are actually the genuine comonomers.     

Inhibition and labeling of enzymes and abzymes by phosphonate diesters

Reactive phosphonate diesters were designed and prepared as inhibitors of serine proteases and esterases. Inactivation of trypsin, chymotrypsin, and butyrylcholinesterase was determined by residual enzymaticactivity as well as by the release of a chromogenic or fluorogenic product of the inhibition reaction. Second-order rate constants were determined from rates of nitrophenol formation. Application of the reaction for active-site titration of enzyme preparations is demonstrated. A basic functional group present in the nitrophenyl tropane phosphonate diester was shown to confer selectivity for inactivation of try psin and chymotrypsin. Biotinylated derivatives of the phosphonate diesters were prepared to permitanalysis of proteins modified in the inhibition reaction. Labeled polypeptides were resolved by SDSPAGE, electroblotted, and detected by streptavidin-peroxidase staining. A detection limit of less than 4 ng, corresponding to 20 nM of trypsin, was demonstrated. Pretretment of enzymes with DFP or nonbiotinylated phosphonates specifically blocks the labeling. This technique permits identification of serine proteases in complex mixtures with good sensitivity and specificity.     

Generalizing a realization result of B. Zimmermann and K. B. Lee to infranilmanifolds

Let M be a closed infranilmanifold. We show that each finite subgroup G of the outer automorphism group Out ₁(M) admits a finite inflation pHG which can be effectively geometrically realized as a group of affine diffeomorphisms of M. This generalizes an analogous result obtained earlier for closed flat Riemannian manifolds by Lee ([7]) and Zimmermann ([12]). Using the Seifert construction we construct explicitly an example of such a manifold for which Nielsen's realization problem fails. This shows the need for such inflations and can be seen as a complementary example to those in [9].     

Virus Particles Monitored by Fluorescence Spectroscopy: A Potential Detection Assay for Macromolecular Assembly¶

Native fluorescence spectroscopy was used for in situ investigations of two lipid‐containing bacteriophages from the cystovirus family as well as their Pseudomonad host cells. Both the viruses φ6 and φ12 and their bacterial host proteins contain the amino acid tryptophan (trp), which is the predominant fluorophore in UV. Within proteins, trp's structural environment differs, and the differences are reflected in their spectroscopic signatures. It was observed that the peak of the trp emission from both viruses was at 330 nm, a significantly shorter wavelength than trp in either the Pseudomonad host cells or the amino acid's chemical form. This allowed us to monitor the viral attachment process and subsequent lytic release of progeny virus particles by measurement of the trp emission spectra during the infection process. This work demonstrates that fluorescence may offer a novel tool to detect viruses and monitor viral infection of cells and may be part of a biodefense application.