Measurement of acid-catalyzed isomerization of unsaturated aldehyde-2,4-dinitrophenylhydrazone derivatives by high-performance liquid chromatography analysis

Aldehyde-2,4-dinitrophenylhydrazones exist as (E)- and (Z)-geometrical isomers, and adventitious isomerization during sample preparation can cause analytical errors. Purified alkenal-2,4-dinitrophenylhydrazone derivatives comprise only the (E)-isomer. However, partial isomerization to the (Z)-isomer occurs upon addition of acid to attain an equilibrium isomer ratio. The UV-visible spectral properties of the isomers differ; the (Z)-isomer exhibiting a 6-10 nm lower absorption maximum compared to the (E)-isomer. Alkenal-2,4-dinitrophenylhydrazones having a CC double bond at the 2- or 3-position of the alkenal exhibited similar absorption maxima with an equilibrium isomer ratio (0.035) that was much lower than those of other alkenals. The CC double bond at the 3-position migrates to a position of conjugation with the CN double bond during hydrazone synthesis to form a stabilized molecular structure. Alkenal-2,4-dinitrophenylhydrazones having a double bond at the 4-position or greater exhibited a similar absorption maxima equilibrium isomer ratio (0.14) to alkanal-2,4-dinitrophenylhydrazones. The quantitative analysis of carbonyl compounds in air or water using DNPH is usually conducted in the presence of an acid catalyst. Consequently, the solution of the direct extract prepared for HPLC or GC analysis contains both (E)- and (Z)-isomers.     

THE CONFORMATIONAL ANALYSIS AND PHOTOISOMERIZATION OF RETINOCHROME ANALOGS WITH POLYENALS

Abstract— 3, 7-Dimethyl-2, 4, 6, 8, 10-dodecapentaenal was synthesized for reconstitution of the retinochrome analog. Its opsin shift was 1000 cm ¹ smaller than that of native retinochrome, whose chromophore contains the same number of double bonds. The conformational change from 6-s-trans to 6-s-cis , as figured in a retinal molecule, plays an important role in the formation of the retinochrome analog, based on the estimation of opsin shifts for retinal analogs locked in the 6-s conformation. Thus the conformation of the 6–7 single bond in the native retinochrome was suggested to be 6 -cis . Analysis of the circular dichroic spectra of retinochrome analogs revealed that the 6-s conformation is independent of the appearance of the β-band. The stereoselectivity in the photoisomerization of the retinal analogs by a retinochrome template depends on the hydrophobic binding in the region of the β-ionone ring.     

New formation of a 1,3-dipole in the reaction of adamantanone azine with triazolinediones: In connection with reactivity toward singlet oxygen

Tricyclo[3.3.1.1]decanone azine ( adamantanone azine ) reacts with 4-substituted 1,2,4-triazoline-3,5-diones under elimination of diazo adamantane to give a 1,3-dipole ( “azomethinimine” ), which on treatment with dipolahophiles affords the [ 2 + 3] -cycloadducts. The reaction is compared with singlet oxygen oxidation.     

Relation between location of elements in periodic table and affinity for the malignant tumor (author's transl)

Affinity of many inorganic compounds for the malignant tumor was examined, using the rats which were subcutaneously transplanted with Yoshida sarcoma. And the relations between the uptake rate into the malignant tumor and in vitro binding power to the protein were investigated in these compounds. In these experiments, the bipositive ions and anions had not affinity for the tumor tissue with a few exceptions. On the other hand, Hg, Au and Bi, which have strong binding power to the protein, showed high uptake rate into the malignant tumor. As Hg++, Au+ and Bi+++ are soft acids according to classification of Lewis acids, it was thought that these elements would bind strongly to soft base (R-SH, R-S-) present in the tumor tissue. In many hard acids (according to classification of Lewis acids), the uptake rate into the tumor was shown as a function of ionic potentials (valency/ionic radii) of the metal ions. It is presumed that the chemical bond of these hard acids in the tumor tissue is ionic bond to hard base (R-COO-, R-PO3(2-), R-SO3-, R-NH2).     

A theoretical study of the dielectric solvent effect on vicinal spin coupling constants

The dielectric solvent effect on HCCH, HCNH and HCNC spin coupling constants in ethane, tetrachloroethane and trans N-methylformamide has been calculated by finite perturbation theory based on the INDO and CNDO/2 approximations incorporating solvaton theory. The available experimental data are interpreted using the calculated variations of spin coupling constants. The effect of dielectric constant on the general form of the Karplus relation is included in the finite perturbation calculations.     

In Situ Processing of Nano Crystalline Oxide Particles/Polymer Hybrid

Nano sized crystalline particles/polymer hybrids were synthesized form designed metal-organic precursors. The newly developed method is composed of the synthesis of organic matrix by polymerization and the in situ nucleation and growth of crystalline oxide particles in the organic matrix below 100°C. The design of metal-organic precursor modified with polymerizable ligand and the selection of reaction conditions does influence the size and crystallinity of ceramic particles in organic matrix. The nano-sized magnetic particle/polymer hybrid exhibits the interesting feature of superparamagnetism and quantum size effect. The crystalline particles of BaTiO₃/, PbTiO₃/, and KNbO₃/polymer hybrids behave to be dielectric and show the typical electro-rheological behavior.     

Development of rectangular column structured titanium oxide photocatalysts anchored on silica sheets by a wet process

Commercially available powdered photocatalysts such as P-25 are known to show high photocatalytic performance. However, in practical use, their anchoring onto a substrate without any binders is still very difficult. The purpose of this study is to design and develop high-performance photocatalysts that can be anchored onto a substrate, and to this end we have prepared a titanium oxide photocatalyst using a wet process. The results of this study led to the successful development of rectangular column structured titanium oxide crystals which could be anchored onto silica sheets. The rectangular columnar crystal was 100- 500 nm wide and 1000- 5000 nm long. Moreover, investigations on the complete oxidation reaction of acetaldehyde into CO₂ and H₂O showed a high performance equivalent to that of the most efficient marketed powdered photocatalysts.     

Electrophoretic analysis of phosphorylation of the yeast 20S proteasome

The 26S proteasome complex, consisting of two multisubunit complexes, a 20S proteasome and a pair of 19S regulatory particles, plays a major role in the nonlysosomal degradation of intracellular proteins. The 20S proteasome was purified from yeast and separated by two-dimensional gel electrophoresis (2-DE). A total of 18 spots separated by 2-DE were identified as the 20S proteasome subunits by peptide mass fingerprinting with matrix assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS). The alpha2-, alpha4- and alpha7-subunits gave multiple spots, which converged into one spot for each subunit when treated with alkaline phosphatase. The difference of pI between phosphorylated and dephosphorylated spots and their reaction against anti-phosphotyrosine antibody suggested that the alpha2- and alpha4-subunits are phosphorylated either at Ser or at Thr residue, and the alpha7-subunit is phosphorylated at Tyr residue(s). These phosphorylated subunits were analyzed by electrospray ionization-quadrupole time of flight-tandem MS (ESI-QTOF-MS/MS) to deduce the phosphorylation sites. The 20S proteasome has three different protease activities: chymotrypsin-like, trypsin-like and peptidylglutamyl peptide-hydrolyzing activities. The phosphatase treatment increased K(m) value for chymotrypsin-like activity of the 20S proteasome, indicating that phosphorylation may play an important role in regulating the proteasome activity.     

Automated multi-attribute method sample preparation using high-throughput buffer exchange tips

Rationale

The multi-attribute method (MAM) has become a valuable mass spectrometry (MS)-based tool that can identify and quantify the site-specific product attributes and purity information for biotherapeutics such as monoclonal antibodies (mAbs) and fusion molecules in recent years. As we expand the use of the MAM at various stages of drug development, it is critical to enhance the sample preparation throughput without additional chemical modifications and variability.

Methods

In this study, a fully automated MAM sample preparation protocol is presented, where rapid desalting in less than 15 minutes is achieved using miniaturized size-exclusion chromatography columns in pipette tips on an automated liquid handler. The peptide samples were analyzed using an electrospray ionization (ESI) orbitrap mass spectrometer coupled to an ultra-high-performance liquid chromatography (UHPLC) system with a dual column switching system.

Results

No significant change was observed in product attributes and their quantities compared with manual, low-artifact sample preparation. The sample recovery using the buffer exchange tips was greatly enhanced over the manual spin cartridges while still demonstrating excellent reproducibility for a wide variety of starting sample concentrations. Unlike a plate desalting system, the individual columns provide flexibility in the number of samples prepared at a time and sample locations within plates.

Conclusions

This automated protocol enables the preparation of up to 96 samples with less “at-bench” time than the manual preparation of a smaller batch of samples, thereby greatly facilitating support of process development and the use of the MAM in quality control.

     

A New Family of Indoaniline-Derived Calix[4]arenes: Synthesis and Optical Recognition Properties as a Chromogenic Receptor(1)

A new family of indoaniline-derived calix[4]arenes has been synthesized for the purpose of developing a new chromogenic receptor. A condensing reaction of calix[4]arene (1) with 4-(diethylamino)-2-methylaniline hydrochloride (2) in the presence of an oxidizing agent under alkaline conditions affords mono- (3), 1,2-bis- (4), 1,3-bis- (5), and tetrakisindoaniline-derived (6) calix[4]arenes after careful column chromatography. Compound 3 is crystallized from a CHCl(3)-MeOH solution, and the crystal structure was determined by X-ray analysis. The crystal is monoclinic, space group P2(1)/n, Z = 4, a = 19.507(6) ?, b = 18.591(6) ?, c = 8.524(2) ?, beta = 94.69(2) degrees. The final R value for 2406 reflections of F(o) > 3sigma(F(o)) is 0.085. A unique intramolecular hydrogen-bonding network involving the carbonyl oxygen of indoaniline for 3 implied that the quinone carbonyl group as an acceptor of the chromophore can easily be subjected to an electrostatic interaction in the lower rim. Indeed, 1,3-bis(indoaniline)-derived 2,4-bis((ethoxycarbonyl)methoxy)calix[4]arene 7, prepared by the reaction of 5 with ethyl bromoacetate in the presence of NaH, is capable of undergoing an efficient ion-dipole interaction between the binding cation and the two quinone carbonyl groups of the chromophores, so that a selective Ca(2+)-induced pronounced color change (wavelength change > 100 nm) occurs with an association constant on the order of 10(6) in 99% EtOH, making 7 of potential use as an optical sensor for Ca(2+) detection. The IR and NMR studies have indicated that Ca(2+) is encapsulated in the cavity made by the distally located OCH(2)CO(2) groups on the lower rim of the cone-shaped calix[4]arene segment. Interestingly, however, the shape of the cavity in which Ca(2+) has been encapsulated does not have a C(2) axis of symmetry, as inferred from the (1)H-(1)H COSY experiment. On the other hand, 1,2-bis(indoaniline)-derived analogue 8 shows no response with metal ions, which can be interpreted to mean the absence of a cavity for encapsulation on the lower rim.