Optimization of Insecticidal Triterpene Derivatives by Biomimetic Oxidations with Hydrogen Peroxide and Iodosobenzene Catalyzed by MnIII and FeIII Porphyrin Complexes

Semisynthetic functionalized triterpenes (4α,14‐dimethyl‐5α,8α‐8,9‐epoxycholestan‐3β‐yl acetate; 4α,14‐dimethyl‐5α‐cholest‐8‐ene‐3,7,11‐trione; 4α,14‐dimethyl‐5α‐cholesta‐7,9(11)‐dien‐3‐one and 4α,14‐dimethyl‐5α‐cholest‐8‐en‐3β‐yl acetate), previously prepared from 31‐norlanostenol, a natural insecticide isolated from the latex of Euphorbia officinarum, have been subjected to oxidation with hydrogen peroxide (H₂O₂) and iodosobenzene (PhIO) catalyzed by porphyrin complexes (cytochrome P‐450 models) in order to obtain optimized derivatives with high regioselectivity. The main transformations were epoxidation of the double bonds and hydroxylations of non‐activated C–H groups and the reaction products were 25‐hydroxy‐4α,14‐dimethyl‐5α‐cholesta‐7,9(11)‐dien‐3β‐yl acetate (59 %), 25‐hydroxy‐4α,14‐dimethyl‐5α‐cholest‐8‐ene‐3,7,11‐trione (60 %), 4α,14‐dimethyl‐5α,7β‐7,8‐epoxycholest‐9(11)‐en‐3‐one (22 %), 8‐hydroxy‐4α,14‐dimethyl‐5α‐cholest‐9(11)‐ene‐3,7‐dione (16 %), 12α‐hydroxy‐4α,14‐dimethyl‐5α,7β‐7,8‐epoxycholest‐9(11)‐en‐3‐one (16 %), and 4α,14‐dimethyl‐5α,8α‐8,9‐epoxycholestan‐3β‐yl acetate (26 %), respectively. We also investigated the insect (Myzus persicae, Rhopalosiphum padi and Spodoptera littoralis) antifeedant and postingestive effects of these terpenoid derivatives. None of the compounds tested had significant antifeedant effects, however, all were more effective postingestive toxicants on S. littoralis larvae than the natural compound 31‐norlanostenol, with 4α,14‐dimethyl‐5α,8α‐8,9‐epoxycholestan‐3β‐yl acetate being the most active. The study of their structure–activity relationships points out at the importance of C3 and C7 substituents.     

Structural investigation of the complexation properties between horse spleen apoferritin and metalloporphyrins

Crystallographic studies of L-chain horse spleen apoferritin (HSF) co-crystallized with Pt-hematoporphyrin IX and Sn-protoporphyrin IX have brought significant new insights into structure-function relationships in ferritins. Interactions of HSF with porphyrins are discussed. Structural results show that the nestling properties into HSF are dependent on the porphyrin moiety. (Only protoporphyrin IX significantly interacts with the protein, whereas hematoporphyrin IX does not.) These studies additionally point out the L-chain HSF ability to demetalate metalloporphyrins, a result which is of importance in looking at the iron storage properties of ferritins. In both compound investigated (whether the porphyrin reaches the binding site or not), the complexation appears to be concomitant with the extraction of the metal from the porphyrin. To analyze further the previous results, a three-dimensional alignment of ferritin sequences based on available crystallographic coordinates, including the present structures, is given. It confirms a high degree of homology between these members of the ferritin family and thus allows us to emphasize observed structural differences: 1) unlike L-chain HSF, H-chain human ferritin presents no preformed binding site; and 2) despite the absence of axial ligands, and due to the demetalation, L-chain HSF is able to host protoporphyrin at a similar location to that naturally found in bacterioferritin.     

Determination of the absolute configuration of Anisotome irregular diterpenes: application of CD and NMR methods

Several Anisotome diterpene derivatives were synthesized in an attempt to obtain a crystalline compound for X-ray analysis. Although we were unable to obtain a suitable crystal, the absolute configuration of the irregular diterpene skeleton was determined using two other techniques: a circular dichroism (CD) protocol based on a tetraarylporphyrin molecular tweezer that allowed prediction of the absolute stereochemistry on a microscale level, and a method employing differences in NMR shifts from derivatization of the naturally occurring acid 1 with enantiomers of a phenylglycine methyl ester (PGME) chiral anisotropic reagent. The excellent agreement between the CD and NMR methods led to the assignment of a 2S-absolute configuration for anisotomenoic acid 1.     

Iron porphyrin treatment extends survival in a transgenic animal model of amyotrophic lateral sclerosis

Oxidative damage, produced by mutant Cu/Zn superoxide dismutase (SOD1), may play a role in the pathogenesis of amyotrophic lateral sclerosis (ALS), a devastating motor neuron degenerative disease. A novel approach to antioxidant therapy is the use of metalloporphyrins that catalytically scavenge a wide range of reactive oxygen and reactive nitrogen species. In this study, we examined the therapeutic potential of iron porphyrin (FeTCPP) in the G93A mutant SOD1 transgenic mouse model of ALS. We found that intraperitoneal injection of FeTCPP significantly improved motor function and extended survival in G93A mice. Similar results were seen with a second group of mice wherein treatment with FeTCPP was initiated at the onset of hindlimb weakness-roughly equivalent to the time at which treatment would begin in human patients. FeTCPP-treated mice also showed a significant reduction in levels of malondialdehyde (a marker of lipid peroxidation), in total content of protein carbonyls (a marker of protein oxidation), and increased neuronal survival in the spinal cord. These results therefore provide further evidence of oxidative damage in a mouse model of ALS, and suggest that FeTCPP could be beneficial for the treatment of ALS patients.     

Gravity perception requires statoliths settled on specific plasma membrane areas in characean rhizoids and protonemata

Summary. The noninvasive infrared laser micromanipulation technique (optical tweezers, optical trapping) and centrifugation were used to study susception and perception, the early events in the gravitropic pathway of tip-growing characean rhizoids and protonemata. Reorientation of the growth direction in both cell types was only initiated when at least 2–3 statoliths settled on specific areas of the plasma membrane. This statolith-sensitive plasma membrane area is confined to the statolith region (10–35 μm behind the tip) in positively gravitropic rhizoids, whereas in negatively gravitropic protonemata, this area is limited to the apical plasma membrane (0–10 μm). Statolith sedimentation towards the sensitive plasma membrane areas is mediated by the concerted action of actin and gravity. The process of sedimentation, the pure physical movement, of statoliths is not sufficient to initiate graviresponses in both cell types. It is concluded that specific statolith-sensitive plasma membrane areas play a crucial role in the signal transduction pathway of gravitropism. These areas may represent the primary sites for gravity perception and may transform the information derived from the gravity-induced statolith sedimentation into physiological signals which trigger the molecular mechanisms of the opposite graviresponses in characean rhizoids and protonemata. Received September 10, 2001 Accepted November 16, 2001     

Serine phosphorylation on position 1033 of vinculin impacts cellular mechanics

This study evaluates the influence of S1033 vinculin phosphorylation on the mechanical properties of cells. We demonstrate that MEFvcl KO cells transfected with the non-phosphorylatable eGFP-vinculin mutant S1033A are of lower stiffness compared to MEFvcl Rescue and phospho-mimicking mutant S1033D cells, which were of similar stiffness. Analogous, 2D traction microscopy indicates that MEFvcl Rescue and MEF mutant S1033D cells generate similar strain energy, but mutant S1033A cells display ∼50% less strain energy. Fluorescence recovery after photobleaching demonstrates that the recovery time for mutant S1033A was significantly lower compared to MEFvcl Rescue and mutant S1033D and that the mobile fraction was smaller for MEFvcl Rescue and mutant S1033D than for mutant S1033A cells. This indicates that serine phosphorylation is required for the activation of vinculin and force transmission in focal adhesions.     

Protective effects of a new metalloporphyrin on paraquat-induced oxidative stress and apoptosis in N27 cells

Paraquat （PQ, 1,1′-dimethyl-4,4′-bipyridinium）,awidely-used herbicide, has been suggested as a potential etiologic factor for the development of Parkinson＇s disease. In recent years, many studies have focused on the mechanism（s） of PQ neurotoxicity. In this study, we examined the neuroprotective effect of manganese （Ⅲ） meso-tetrakis （N,N′-diethylimi- dazolium） porphyrin （MnTDM）, a superoxide dismutase/ catalase mimetic, on PQ-induced oxidative stress and apoptosis in 1 RB3AN27 （N27） cells, a dopaminergic neuronal cell line. The results indicated that MnTDM significantly attenuated PQ-induced loss of cell viability, glutathione depletion, and reactive oxygen species production. MnTDM also ameliorated PQ-induced morphological nuclear changes of apoptosis and increased rates of apoptosis. In addition, our data provide direct evidence that MnTDM suppressed PQ- induced caspase-3 cleavage, possibly a key event of PQ neurotoxicity. These observations suggested that oxidative stress and apoptosis are implicated in PQ-induced neurotoxicity and this toxicity could be prevented by MnTDM. These findings also proposed a novel therapeutic approach for Parkinson＇s disease and other disorders associated with oxidative stress.     

CD‐sensitive Zn‐porphyrin tweezer host‐guest complexes,part 2: Cis‐ and trans‐3‐hydroxy‐4‐aryl/alkyl‐β‐lactams. A case study

This article describes an application of the host‐guest chiral recognition approach called tweezer methodology for the determination of the absolute configuration of 3‐hydroxy‐β‐lactams. These substrates represent challenging cases due to their chemical reactivity, the presence of multiple stereogenic centers and several functional groups which offer various possibilities of binding to the Zn‐porphyrin host. OPLS‐2005, the force field used in this work to predict the interporphyrin twist, modeled correctly the host‐guest complexation mechanism and revealed conformational details of the bound substrates. The computational study also suggested that in cases where an increase in the magnitude of the stereodifferentiation and an intense experimental CD are observed, the bound conformation of the conjugates are hydrogen bonded. The present investigation provides evidence that when the tweezer method is assisted by the OPLS‐2005 based computational approach, it can be successfully applied to the configurational and conformational elucidation of multi‐functional compounds with multiple stereogenic centers. Chirality, 2010. © 2009 Wiley‐Liss, Inc.     

Chiral recognition of cyclic alpha-hydroxyketones by CD-sensitive zinc tetraphenylporphyrin tweezer

A combined chemical/chiroptical microscale protocol for the determination of absolute configurations of cyclic alpha-hydroxyketones is described. The hydroxyl group in cyclic alpha-hydroxyketones is converted into (3-aminopropylamino)acetate (NH2CH2CH2CH2NHCH2COOR), or more generally, according to a newly developed protocol, into (3-hydroxypropylamino)acetate group (HOCH2CH2CH2NHCH2COOR). The resultant conjugated compound forms a 1:1 host-guest complex with a dimeric zinc porphyrin tweezer, which exhibits exciton-coupled bisignate CD spectrum centered around the 420-nm porphyrin Soret band due to induced helicity between the two porphyrins in the complex. The absolute configurations of the alpha-stereogenic center is then determined by comparison of the sign of the observed CD exciton couplet of the complex with that of the preferred porphyrin twist predicted by the Merck Molecular Force Field (MMFFs) method.     

Cell motility in a new single-cell wound model

Summary  Until now researchers have used a monolayer of cultured cells to investigate cell motility toward an injured cell. However, we suspect that, when using this method, adjacent cells move to the free space due to relief of contact inhibition. The current study was designed to investigate the cell motility nearby an injured cell in varying cell connectivity. A lowpower laser beam was used to damage one cell selectively with the silver coating beads. After injury, we observed the cell motility in three different cell types: (1) those immediately adjacent to the injured cell, 92) those removed from the injured cell by interposition of another cell, and (3) those removed from the injured cell by free space. The cells that are in direct contact with the injured cell moved toward the injured cell within 1.5–3.0 h. Indirectly connected cells and cells with no contact, on the other hand, showed no significant movement toward the injured cell. This suggests that the cell motility toward the cell injury is not only due to relief of contact inhibition but might also be caused by cell-to-cell signaling via cell connection. The current method will provide a tool to create a cell injury without damaging adjacent cells.