A formal vinylic substitution reaction for the synthesis of α,ß-unsaturated enol esters and their anticancer potential.

Arylsulfonyl group-bearing α,ß-unsaturated enol esters were readily assembled via the Cs2CO3-mediated union of 2-bromoallyl sulfones and cinnamic acids. The overall transformation is equivalent to an sp2 carbon-oxygen coupling reaction, and therefore constitutes a formal vinylic substitution. Several of the products display promising levels of antiproliferative activities higher than that of the anticancer drug carboplatin. Thiophenol reacted with 2-bromoallyl sulfones under identical conditions to afford α-thiophenyl-α'-tosyl acetone via an apparent aerial oxidation.

Understanding the Influence of API Conformations on Amorphous Dispersion Formation Potential Predictions using the R3m Molecular Descriptor.

The R3m molecular descriptor (R-GETAWAY third-order autocorrelation index weighted by the atomic mass) has previously been shown to encode molecular attributes that appear to be physically and chemically relevant to grouping diverse active pharmaceutical ingredients (API) according to their potential to form persistent amorphous solid dispersions (ASDs) with polyvinylpyrrolidone-vinyl acetate copolymer (PVPVA). The initial R3m dispersibility model was built by using a single three-dimensional (3D) conformation for each drug molecule. Since molecules in the amorphous state will adopt a distribution of conformations, molecular dynamics simulations were performed to sample conformations that are probable in the amorphous form, which resulted in a distribution of R3m values for each API. Although different conformations displayed R3m values that differed by as much as 0.4, the median of each R3m distribution and the value predicted from the single 3D conformation were very similar for most structures studied. The variability in R3m resulting from the distribution of conformations was incorporated into a logistic regression model for the prediction of ASD formation in PVPVA, which resulted in a refinement of the classification boundary relative to the model that only incorporated a single conformation of each API.

Fluorescent polymer as a biosensing tool for the diagnosis of microbial pathogens.

Diseases and diagnoses are predominant in the human population. Early diagnosis of etiological agents plays a vital role in the treatment of bacterial infections. Existing standard diagnostic platforms are laborious, time-consuming, and require trained personnel and cost-effective procedure, though they are producing promising results. These shortcomings have led to a thirst for rapid diagnostic procedures. Fluorescence-based diagnosis is one of the efficient rapid diagnostic methods that rely on specific and sensitive bacterial detection. Emerging bio-sensing studies on conducting polymers (CPs) are gaining popularity in medical diagnostics due to their promising properties of high fluorescence efficiency, good light stability, and low cytotoxicity. Poly[2-methoxy-5-(2'-ethylhexyloxy)-1,4-phenylenevinylene] (MEH-PPV), is the first identified soluble polymer and model material for understanding the fundamental photophysics of conventional CPs. In this present study, MEH-PPV is used as a fluorescent dye for direct pathogen detection applications by interacting with the microbial cell surface. An optimized concentration of MEH-PPV solution used to confirm the presence of selective bacterial structures. The present study endeavours towards bacterial detection based on the emission from bacteria due to interfacial interaction between polymer and bacterial surface.

Dissolution Mechanisms of Amorphous Solid Dispersions: A Close Look at the Dissolution Interface.

Despite the recent success of amorphous solid dispersions (ASDs) at enabling the delivery of poorly soluble small molecule drugs, ASD-based dosage forms are limited by low drug loading. This is partially due to a sharp decline in drug release from the ASD at drug loadings surpassing the 'limit of congruency' (LoC). In some cases, the LoC is as low as 5% drug loading, significantly increasing the risk of pill burden. Despite efforts to understand the mechanism responsible for the LoC, a clear picture of the molecular processes occurring at the ASD/solution interface remains elusive. In this study, the ASD/solution interface was studied for two model compounds formulated as ASDs with copovidone. The evolution of a gel layer and its phase behavior was captured in situ with fluorescence confocal microscopy, where fluorescent probes were added to label the hydrophobic and hydrophilic phases. Phase separation was detected in the gel layer for most of the ASDs. The morphology of the hydrophobic phase was found to correlate with the release behavior, where a discrete phase resulted in good release and a continuous phase formed a barrier leading to poor release. The continuous phase formed at a lower drug loading for the system with stronger drug-polymer interactions. This was due to incorporation of the polymer into the hydrophobic phase. The study highlights the complex molecular and phase behavior at the ASD/solution interface of copovidone-based ASDs and provides a thermodynamic argument for qualitatively predicting the release behavior based on drug-polymer interactions.

Nano-liquid chromatography with monolithic stationary phase based on naphthyl monomer for proteomics analysis.

Monolithic poly(2-vinylnaphthalene-co-divinylbenzene) columns were introduced, for the first time, and were evaluated as the separation media for nano-liquid chromatography (nano-LC). These columns were prepared by in-situ polymerization of 2-vinylnaphthalene (2-VNA) as the functional monomer and divinylbenzene (DVB) as the crosslinker in a fused silica capillary column of 50 µm i.d. Various porogenic solvents, including tetrahydrofuran (THF), dodecanol and toluene were used for morphology optimization. Final monolithic column (referred to as VNA column) was characterized by using scanning electron microscopy (SEM) and chromatographic analyses. Alkylbenzenes (ABs), and polyaromatic hydrocarbons (PAHs) were separated using the VNA column while the column offered excellent hydrophobic and π-π interactions under reversed-phase conditions. Theoretical plates number up to 41,200 plates/m in isocratic mode for ethylbenzene could be achieved. The potential of the final VNA column was demonstrated with a gradient elution in the  separation of six intact proteins, including ribonuclease A (RNase A), cytochrome C (Cyt C), lysozyme (Lys), ß-lactoglobulin (ß-lac), myoglobin (My) and α-chymotrypsinogen (α-chym) in nano LC system. The column was then applied to the peptide analysis of trypsin digested cytochrome C, allowing a high peak capacity up to 1440 and the further proteomics analysis of COS-7 cell line was attempted applying the final monolithic column in nano-LC UV system.

Release Mechanisms of Amorphous Solid Dispersions: Role of Drug-Polymer Phase Separation and Morphology.

Formulating poorly soluble molecules as amorphous solid dispersions (ASDs) is an effective strategy to improve drug release. However, drug release rate and extent tend to rapidly diminish with increasing drug loading (DL). The poor release at high DLs has been postulated to be linked to the process of amorphous-amorphous phase separation (AAPS), although the exact connection between phase separation and release properties remains somewhat unclear. Herein, release profiles of ASDs formulated with ritonavir (RTV) and polyvinylpyrrolidone/vinyl acetate (PVPVA) at different DLs were determined using surface normalized dissolution. Surface morphologies of partially dissolved ASD compacts were evaluated with confocal fluorescence microscopy, using Nile red and Alexa Fluor 488 as fluorescence markers to track the hydrophobic and hydrophilic phases respectively. ASD phase behavior during hydration and release of components were also visualized in real time using a newly developed in situ confocal fluorescence microscopy method. RTV-PVPVA ASDs showed complete and rapid drug release below 30% DL, partial drug release at 30% DL and no drug release above 30% DL. It was observed that formation of discrete drug-rich droplets at lower DLs led to rapid and congruent release of both drug and polymer, whereas formation of continuous drug-rich phase at the ASD matrix-solution interface was the cause of poor release above certain DLs. Thus, the domain size and interconnectivity of phase separated drug-rich domains appear to be critical factors impacting drug release from RTV-PVPVPA ASDs.

The impact of applying an additional polymer coating on high drug-loaded amorphous solid dispersions layered onto pellets.

Inhibiting surface crystallization is an interesting strategy to enhance the physical stability of amorphous solid dispersions (ASDs), still preserving high drug loads. The aim of this study was to investigate the potential surface crystallization inhibitory effect of an additional polymer coating onto ASDs, comprising high drug loads of a fast crystallizing drug, layered onto pellets. For this purpose, bilayer coated pellets were generated with fluid-bed coating, of which the first layer constitutes a solid dispersion of naproxen (NAP) in poly(vinylpyrrolidone-co-vinyl acetate) (PVP-VA) in a 40:60 or 35:65 (w/w) ratio, and ethyl cellulose (EC) composes the second layer. The physical stability of these double-layered pellets, in comparison to pellets with an ASD layer only, was assessed under accelerated conditions by monitoring with X-ray powder diffraction (XRPD) at regular time intervals. Bilayer coated pellets were however found to be physically less stable than pellets with an ASD layer only. Applying the supplementary EC coating layer induced crystallization and heterogeneity in the 40:60 and 35:65 (w/w) NAP-PVP-VA ASDs, respectively, attributed to the initial contact with the solvent. Caution is thus required when applying an additional coating layer on top of an ASD layer with fluid-bed coating, for instance for controlled release purposes, especially if the ASD consists of high loads of a fast crystallizing drug.

Canolol and its derivatives: A novel bioactive with antioxidant and anticancer properties.

The health and safety concerns associated with synthetic antioxidants has resulted in an urgent search for natural sources of antioxidants. Such antioxidants are not only convenient but may also have important therapeutic properties. Oilseed crops, for example, are rich in phenolic compounds some of which exhibit powerful antioxidant properties that have broad applications in both the food and feed industry. Often, the concentration of these phenolic compounds is affected by many processing conditions including temperature, pressure, pH, and extracting solvents. Hence it is important to optimize processing conditions to obtain maximum levels of those antioxidants with superior antioxidant activity. Oilseeds, such as canola and mustard, are rich sources of sinapates and kaempferol derivatives. When subjected to different processing conditions, including pressurized temperature, sinapates are converted to vinyl phenol derivatives, of which the major one is canolol. This chapter will focus on the nature of canolol and its applications in food and medicine.

Cleavable Comonomers for Chemically Recyclable Polystyrene: A General Approach to Vinyl Polymer Circularity.

Many common polymers, especially vinyl polymers, are inherently difficult to chemically recycle and are environmentally persistent. The introduction of low levels of cleavable comonomer additives into existing vinyl polymerization processes could facilitate the production of chemically deconstructable and recyclable variants with otherwise equivalent properties. Here, we report thionolactones that serve as cleavable comonomer additives for the chemical deconstruction and recycling of vinyl polymers prepared through free radical polymerization, using polystyrene (PS) as a model example. Deconstructable PS of different molar masses (∼20-300 kDa) bearing varied amounts of statistically incorporated thioester backbone linkages (2.5-55 mol %) can be selectively depolymerized to yield well-defined thiol-terminated fragments (<10 kDa) that are suitable for oxidative repolymerization to generate recycled PS of nearly identical molar mass to the parent material, in good yields (80-95%). A theoretical model is provided to generalize this molar mass memory effect. Notably, the thermomechanical properties of deconstructable PS bearing 2.5 mol % of cleavable linkages and its recycled product are similar to those of virgin PS. The additives were also shown to be effective for deconstruction of a cross-linked styrenic copolymer and deconstruction and repolymerization of a polyacrylate, suggesting that cleavable comonomers may offer a general approach toward circularity of many vinyl (co)polymers.

Orthogonal C-B Bond Transformation as an Approach for Versatile Synthesis of End-Functionalized Polymers.

Conventionally inaccessible end-functionalized vinyl polymers were synthesized via orthogonal side-chain replacement for terminal and repeating units of poly(alkenyl boronate)s. A terminal-defined polymer of isopropenyl boronic acid pinacol ester (IPBpin) was synthesized via RAFT polymerization, and subsequent cobalt (Co)-catalyzed end olefination afforded the polymer carrying the C(sp2)-B bond at the terminal and the C(sp3)-B bond in repeating units. Herein, the terminal C(sp2)-B bond was selectively transformable via palladium (Pd)-catalyzed Suzuki-Miyaura cross coupling, and subsequent transformation of the repeating C(sp3)-B unit gave the poly(α-methyl vinyl alcohol) [poly(MVA)] bearing various functional groups at the terminal. The boron-based stepwise polymer reaction thus overcame the synthetic difficulty of the end-functionalized poly(MVA), which is ascribed to the poor polymerization ability of the corresponding acetate monomer, i.e., isopropenyl acetate.

Synthesis of Crosslinked 2'-OMe RNA Duplexes Using 2-Amino-6-Vinylpurine and Their Application for Effective Inhibition of miRNA Function.

Crosslinking reactions to nucleic acids are an effective way to prepare stable complexes formed by covalent bonding. We demonstrated that fully 2'-O-methylated (2'-OMe) RNAs having a 2-amino-6-vinylpurine (AVP) exhibited an efficient crosslinking to uracil in the target RNA. Recently, we reported the preparation of crosslinked 2'-OMe RNA duplexes using AVP and the anti-miRNA oligonucleotides (AMOs) containing crosslinked duplexes at the terminal positions. These AMOs exhibited efficient microRNA (miRNA) inhibition at very low concentrations. In this article, we describe the chemical synthesis of 2'-OMe oligonucleotides containing AVP and preparation of the AMOs bearing crosslinked 2'-OMe RNA duplexes using AVP. In addition, we describe in detail the miRNA inhibition assay using these AMOs. © 2022 Wiley Periodicals LLC. Basic Protocol 1: Synthesis of phosphoramidite of 2-amino-6-vinylguanosine derivative Basic Protocol 2: Synthesis of AVP-2'-OMe RNA Basic Protocol 3: Evaluation of the crosslink reactivity of CFO containing AVP to the 2'-OMe RNA and preparation of AMOs containing crosslinked duplex Basic Protocol 4: miRNA inhibition assays.

Role of Surfactants on Release Performance of Amorphous Solid Dispersions of Ritonavir and Copovidone.

PURPOSE: To understand the role of different surfactants, incorporated into amorphous solid dispersions (ASDs) of ritonavir and copovidone, in terms of their impact on release, phase behavior and stabilization of amorphous precipitates formed following drug release. METHODS: Ternary ASDs with ritonavir, copovidone and surfactants (30:70:5 w/w/w) were prepared by rotary evaporation. ASD release performance was tested using Wood's intrinsic dissolution rate apparatus and compared to the binary drug-polymer ASD with 30% drug loading. Size measurement of amorphous droplets was performed using dynamic light scattering. Solid state characterization was performed using attenuated total reflectance-infrared spectroscopy, differential scanning calorimetry and scanning electron microscopy. RESULTS: All surfactant-containing ASDs showed improvement over the binary ASD. Span 85 and D-α-tocopheryl polyethylene glycol succinate (TPGS) showed complete release with no evidence of AAPS or crystallization whereas Span 20 and Tween 80 showed < 50% release with amorphous amorphous phase separation (AAPS). Span 20 also induced solution crystallization. Sodium dodecyl sulfate (SDS) showed very rapid, albeit incomplete (~ 80%) release. AAPS was not observed with SDS. However, crystallization on the dissolving solid surface was noted. Span 20 and TPGS formed the smallest and most size-stable droplets with ~ 1 µm size whereas coalescence was noted with other surfactants. CONCLUSIONS: Surfactants improved the release performance relative to the binary ASD. Different surfactant types impacted overall performance to varying extents and affected different attributes. Overall, Span 85 showed best performance (complete release, no crystallization/AAPS and small droplet size). Correlation between physicochemical properties and surfactant performance was not observed.

Each phospholipase A2 type exhibits distinct selectivity toward sn-1 ester, alkyl ether, and vinyl ether phospholipids.

Glycerophospholipids are major components of cell membranes and have enormous variation in the composition of fatty acyl chains esterified on the sn-1 and sn-2 position as well as the polar head groups on the sn-3 position of the glycerol backbone. Phospholipase A2 (PLA2) enzymes constitute a superfamily of enzymes which play a critical role in metabolism and signal transduction by hydrolyzing the sn-2 acyl chains of glycerophospholipids. In human cell membranes, in addition to the conventional diester phospholipids, a significant amount is the sn-1 ether-linked phospholipids which play a critical role in numerous biological activities. However, precisely how PLA2s distinguish the sn-1 acyl chain linkage is not understood. In the present study, we expanded the technique of lipidomics to determine the unique in vitro specificity of three major human PLA2s, including Group IVA cytosolic cPLA2, Group VIA calcium-independent iPLA2, and Group V secreted sPLA2 toward the linkage at the sn-1 position. Interestingly, cPLA2 prefers sn-1 vinyl ether phospholipids known as plasmalogens over conventional ester phospholipids and the sn-1 alkyl ether phospholipids. iPLA2 showed similar activity toward vinyl ether and ester phospholipids at the sn-1 position. Surprisingly, sPLA2 preferred ester phospholipids over alkyl and vinyl ether phospholipids. By taking advantage of molecular dynamics simulations, we found that Trp30 in the sPLA2 active site dominates its specificity for diester phospholipids.

Interpreting the Physicochemical Meaning of a Molecular Descriptor Which Is Predictive of Amorphous Solid Dispersion Formation in Polyvinylpyrrolidone Vinyl Acetate.

A molecular descriptor known as R3m (the R-GETAWAY third-order autocorrelation index weighted by the atomic mass) was previously identified as capable of grouping members of an 18-compound library of organic molecules that successfully formed amorphous solid dispersions (ASDs) when co-solidified with the co-polymer polyvinylpyrrolidone vinyl acetate (PVPva) at two concentrations using two preparation methods. To clarify the physical meaning of this descriptor, the R3m calculation is examined in the context of the physicochemical mechanisms of dispersion formation. The R3m equation explicitly captures information about molecular topology, atomic leverage, and molecular geometry, features which might be expected to affect the formation of stabilizing non-covalent interactions with a carrier polymer, as well as the molecular mobility of the active pharmaceutical ingredient (API) molecule. Molecules with larger R3m values tend to have more atoms, especially the heavier ones that form stronger non-covalent interactions, generally, more irregular shapes, and more complicated topology. Accordingly, these molecules are more likely to remain dispersed within PVPva. Furthermore, multiple linear regression modeling of R3m and more interpretable descriptors supported these conclusions. Finally, the utility of the R3m descriptor for predicting the formation of ASDs in PVPva was tested by analyzing the commercially available products that contain amorphous APIs dispersed in the same polymer. All of these analyses support the conclusion that the information about the API geometry, size, shape, and topological connectivity captured by R3m relates to the ability of a molecule to interact with and remain dispersed within an amorphous PVPva matrix.

Heterogenous hydrogel mimicking the osteochondral ECM applied to tissue regeneration.

Inspired by the intricate extracellular matrix (ECM) of natural cartilage and subchondral bone, a heterogenous bilayer hydrogel scaffold is fabricated. Gelatin methacrylate (GelMA) and acryloyl glucosamine (AGA) serve as the main components in the upper layer, mimicking the chondral ECM. Meanwhile, vinylphosphonic acid (VPA) as a non-collagen protein analogue is incorporated into the bottom layer to induce the in situ biomineralization of calcium phosphate. The two heterogenous layers are effectively sutured together by the inter-diffusion between the upper and bottom layer hydrogels, together with chelation between the calcium ions and alginate added to separate layers. The interfacial bonding between the two different layers was thoroughly investigated via rheological measurements. The incorporation of AGA promotes chondrocytes to produce collagen type II and glycosaminoglycans and upregulates the expression of chondrogenesis-related genes. In addition, the minerals induced by VPA facilitate the osteogenesis of bone marrow mesenchymal stem cells (BMSCs). In vivo evaluation confirms the biocompatibility of the scaffold with minor inflammation and confirms the best repair ability of the bilayer hydrogel. This cell-free, cost-effective and efficient hydrogel shows great potential for osteochondral repair and inspires the design of other tissue-engineering scaffolds.

Anti-Markovnikov hydro(amino)alkylation of vinylarenes via photoredox catalysis.

Photoredox catalysis is a powerful means to generate odd-electron species under mild reaction conditions from a wide array of radical precursors. Herein, we present the application of this powerful catalytic manifold to address the hydroalkylation and hydroaminoalkylation of electronically diverse vinylarenes. This reaction allows for generalized alkene hydroalkylation leveraging common alkyl radical precursors, such as organotrifluoroborate salts and carboxylic acids. Furthermore, utilizing easily accessible α-silyl amine reagents or tertiary amines directly, secondary and tertiary amine moieties can be installed onto monoaryl and diaryl alkenes to access valuable products, including γ,γ-diarylamines pharmacophores. Thus, under a unified system, both hydroalkylation and hydroaminoalkylation of alkenes are achieved. The substrate scope is evaluated through 57 examples, the synthetic utility of the method is demonstrated, and preliminary mechanistic insights are presented.

Formation of Multicolor Nanogels Based on Cationic Polyfluorenes and Poly(methyl vinyl ether-alt-maleic monoethyl ester): Potential Use as pH-Responsive Fluorescent Drug Carriers.

In this study, we employed the copolymer poly(methyl vinyl ether-alt-maleic monoethyl ester) (PMVEMA-Es) and three fluorene-based cationic conjugated polyelectrolytes to develop fluorescent nanoparticles with emission in the blue, green and red spectral regions. The size, Zeta Potential, polydispersity, morphology, time-stability and fluorescent properties of these nanoparticles were characterized, as well as the nature of the interaction between both PMVEMA-Es and fluorescent polyelectrolytes. Because PMVEMA-Es contains a carboxylic acid group in its structure, the effects of pH and ionic strength on the nanoparticles were also evaluated, finding that the size is responsive to pH and ionic strength, largely swelling at physiological pH and returning to their initial size at acidic pHs. Thus, the developed fluorescent nanoparticles can be categorized as pH-sensitive fluorescent nanogels, since they possess the properties of both pH-responsive hydrogels and nanoparticulate systems. Doxorubicin (DOX) was used as a model drug to show the capacity of the blue-emitting nanogels to hold drugs in acidic media and release them at physiological pH, from changes in the fluorescence properties of both nanoparticles and DOX. In addition, preliminary studies by super-resolution confocal microscopy were performed, regarding their potential use as image probes.

Structural characterization and antioxidant property of enzymatic-transesterification derivatives of (-)-epigallocatechin-3-O-gallate and vinyl laurate.

(-)-Epigallocatechin-3-O-gallate(EGCG) was enzymatically modified to enhance the lipophilicity and the antioxidant property. The determination of optimal reaction conditions are as follows: Lipase DF "Amano" 15 and acetone were used as catalyst and solvent, respectively. Equal molar of EGCG and vinyl laurate (1:1); lipase addition of 6.0% (w/w of total substrates); reaction temperature of 50°C and reaction time of 96 h, which obtained the conversion rate of EGCG at 80.1%. The structure of EGCG lauroyl derivatives were 5″-O-lauroyl-EGCG, 3″,5″-2-O-lauroyl-EGCG, and 5',3″,5″-3-O-lauroyl-EGCG, identified by high-performance liquid chromatography-mass spectrometry (HPLC-MS) and nuclear magnetic resonance (NMR). Compared with the logP of precursor EGCG (0.69 ± 0.03), the logP of EGCG lauroyl derivatives was 1.37 ± 0.19, 2.27 ± 0.33, and 3.28 ± 0.37, increasing by 0.98, 2.28, and 3.75 times, respectively (p < 0.05), suggesting the grafted fatty acid chains make EGCG derivatives more lipophilic, and the lipid solubility gradually increased as the number of substituents increased. Furthermore, EGCG lauroyl derivatives had excellent lipid oxidation than that of EGCG. The POVs (peroxide values) of soybean oil with mono-, di-, tri-lauroyl EGCG were significantly reduced by 42%, 47%, and 57% than that of EGCG at 21 days, respectively, indicating the antioxidative inhibition of these derivatives decreased with the increase in substituents. This indicates that these derivatives have broad prospects of the antioxidant application while improving their solubility properties in lipophilic environments/high-fat food. Practical Application: The lipophilic esterification reaction of EGCG catalyzed by new catalytic lipase DF "Amano" 15 was carried out in a non-aqueous solvent.Various reaction factors on a higher conversion rate of EGCG lauroyl derivatives were evaluated. The lipophilicity and antioxidant properties of EGCG lauroyl derivatives were much excellent than that of parent EGCG.

Cellulose nanocrystals concentration and oil-water ratio for solid-liquid controllable emulsion polymerization.

Stabilities of cellulose Pickering emulsions are of great importance to utilize them effectively, but influenced by their complex compositions, such as, colloidal particles, oil phases and water phases. In this work, solid-liquid controllable polymerization products could obtain by adjusting cellulose nanocrystals (CNCs) concentration and vinyl acetate (VAc)-water ratio. The emulsions in zone Ӏ (w/o) and II (o/w) of the three-phase diagram were selected for researching. The polymerization emulsions in zone II illustrated the o/w ratio played a more important role than CNCs concentration in the storage stability and practicality of the polymerized emulsion; The polymer in zone Ӏ showed a large number of porous structures. This is an innovative method that different forms of target products are obtained through the guidance of three-phase diagram, which not only broadens the application field, but also applies to other Pickering emulsion systems.

Pseudopeptides with aldehyde or vinylsulfone warheads: Synthesis and antiproteasomal activity.

The comparative study of new proteasome inhibitors based on salicylic acid-modified pseudo-tripeptides terminated with aldehyde or vinylsulfone is presented. We described the synthesis of 11 pairs of pseudopeptides and their properties related to the proteasome inhibition were determined. The effects of integrated amino acids (combinations of leucine, phenylalanine, tryptophan, proline, cyclohexylalanine or norleucine residues) on the activity of the proteasome were investigated. Compounds preferentially inhibited the chymotrypsin ß5-subunit of the proteasome in cell-based assays compared with the ß1- and ß2-subunits, with IC50 values in mid-nanomolar ranges being obtained for the most active members. Our comparative study demonstrated that aldehydes were able to inhibit the proteasome in cells more effectively than vinylsulfones. These results were corroborated by the accumulation of polyubiquitinated proteins in treated cells, GFP accumulation in a reporter cell line and the ability of new compounds to induce apoptotic cell death.