Rational Design,Synthesis, and DNA Binding Properties of Novel Sequence‐Selective Peptidyl Congeners of Ametantrone

Natural and synthetic compounds characterized by an anthraquinone nucleus represent an important class of anti‐neoplastic agents, the mechanism of action of which is related to intercalation into DNA. Ametantrone (AM) is a synthetic 9,10‐anthracenedione bearing two (hydroxyethylamino)ethylamino residues at positions 1 and 4; along with other anthraquinones and anthracyclines, it shares a polycyclic intercalating moiety and charged side chains that stabilize DNA binding. All these drugs elicit adverse side effects, which represent a challenge for antitumor chemotherapy. In the present work the structure of AM was augmented with appropriate groups that target well‐defined base pairs in the major groove. These should endow AM with DNA sequence selectivity. We describe the rationale for the synthesis and the evaluation of activity of a new series of compounds in which the planar anthraquinone is conjugated at positions 1 and 4 through the side chains of AM or other bioisosteric linkers to appropriate dipeptides. The designed novel AM derivatives were shown to selectively stabilize two oligonucleotide duplexes that both have a palindromic GC‐rich hexanucleotide core, but their stabilizing effects on a random DNA sequence was negligible. In the case of the most effective compound, the 1,4‐bis‐[Gly‐(L ‐Lys)] derivative of AM, the experimental results confirm the predictions of earlier theoretical computations. In contrast, AM had equal stabilizing effects on all three sequences and showed no preferential binding. This novel peptide derivative can be classified as a strong binder regarding the sequences that it selectively targets, possibly opening the exploitation of less cytotoxic conjugates of AM to the targeted treatment of oncological and viral diseases.     

Synthesis,In Vitro,and Computational Studies of PTP1B Phosphatase Inhibitors Based on Oxovanadium(IV) and Dioxovanadium(V) Complexes

One of the main goals of recent bioinorganic chemistry studies has been to design and synthesize novel substances to treat human diseases. The promising compounds are metal-based and metal ion binding components such as vanadium-based compounds. The potential anticancer action of vanadium-based compounds is one of area of investigation in this field. In this study, we present five oxovanadium(IV) and dioxovanadium(V) complexes as potential PTP1B inhibitors with anticancer activity against the MCF-7 breast cancer cell line, the triple negative MDA-MB-231 breast cancer cell line, and the human keratinocyte HaCaT cell line. We observed that all tested compounds were effective inhibitors of PTP1B, which correlates with anticancer activity. [VO(dipic)(dmbipy)]·2 H₂O (Compound 4) and [VOO(dipic)](2-phepyH)·H₂O (Compound 5) possessed the greatest inhibitory effect, with IC₅₀ 185.4 ± 9.8 and 167.2 ± 8.0 nM, respectively. To obtain a better understanding of the relationship between the structure of the examined compounds and their activity, we performed a computer simulation of their binding inside the active site of PTP1B. We observed a stronger binding of complexes containing dipicolinic acid with PTP1B. Based on our simulations, we suggested that the studied complexes exert their activity by stabilizing the WPD-loop in an open position and limiting access to the P-loop.     

Computational modeling of biodegradable blends of starch amylose and poly-propylene carbonate

The current study investigates the nature of interactions between starch (amylose portion) and poly-propylene carbonate by employing DFT based B3LYP and semi-empirical AM1 and PM3 methods on five complexes. The computed negative binding energies confirm the stability of the complexes. Favorable interactions between amylose and poly-propylene carbonate were indicated by the formation of hydrogen bonds. The number of hydrogen bonds identified per poly-propylene carbonate monomer attains a saturation value of 0.57 for AM1 and 0.51 for PM3 for larger complexes. The average binding energy per hydrogen bond was computed to be −12.76 kJ/mol for AM1 and −9.79 kJ/mol for PM3 method. The decrease in the computed vibrational frequencies of the hydroxyl OH and carbonyl CO frequencies of the complexes agrees qualitatively with experimental results, further confirming the presence of favorable interactions. Molecular modeling was thus successful in providing useful insight into the nature of interactions between starch and PPC.     

Negative Cooperativity in NAD(P)H Quinone Oxidoreductase 1 (NQO1)

NAD(P)H quinone oxidoreductase-1 (NQO1) is a homodimeric protein that acts as a detoxifying enzyme or as a chaperone protein. Dicourmarol interacts with NQO1 at the NAD(P)H binding site and can both inhibit enzyme activity and modulate the interaction of NQO1 with other proteins. We show that the binding of dicoumarol and related compounds to NQO1 generates negative cooperativity between the monomers. This does not occur in the presence of the reducing cofactor, NAD(P)H, alone. Alteration of Gly150 (but not Gly149 or Gly174) abolished the dicoumarol-induced negative cooperativity. Analysis of the dynamics of NQO1 with the Gaussian network model indicates a high degree of collective motion by monomers and domains within NQO1. Ligand binding is predicted to alter NQO1 dynamics both proximal to the ligand binding site and remotely, close to the second binding site. Thus, drug-induced modulation of protein motion might contribute to the biological effects of putative inhibitors of NQO1.     

Redox Properties of 3-Iodothyronamine (T1AM) and 3-Iodothyroacetic Acid (TA1)

3-iodothyronamine (T1AM) and 3-iodothyroacetic acid (TA1) are thyroid-hormone-related compounds endowed with pharmacological activity through mechanisms that remain elusive. Some evidence suggests that they may have redox features. We assessed the chemical activity of T1AM and TA1 at pro-oxidant conditions. Further, in the cell model consisting of brown adipocytes (BAs) differentiated for 6 days in the absence (M cells) or in the presence of 20 nM T1AM (M + T1AM cells), characterized by pro-oxidant metabolism, or TA1 (M + TA1 cells), we investigated the expression/activity levels of pro- and anti-oxidant proteins, including UCP-1, sirtuin-1 (SIRT1), mitochondrial monoamine (MAO-A and MAO-B), semicarbazide-sensitive amine oxidase (SSAO), and reactive oxygen species (ROS)-dependent lipoperoxidation. T1AM and TA1 showed in-vitro antioxidant and superoxide scavenging properties, while only TA1 acted as a hydroxyl radical scavenger. M + T1AM cells showed higher lipoperoxidation levels and reduced SIRT1 expression and activity, similar MAO-A, but higher MAO-B activity in terms of M cells. Instead, the M + TA1 cells exhibited increased levels of SIRT1 protein and activity and significantly lower UCP-1, MAO-A, MAO-B, and SSAO in comparison with the M cells, and did not show signs of lipoperoxidation. Our results suggest that SIRT1 is the mediator of T1AM and TA1 pro-or anti-oxidant effects as a result of ROS intracellular levels, including the hydroxyl radical. Here, we provide evidence indicating that T1AM and TA1 administration impacts on the redox status of a biological system, a feature that indicates the novel mechanism of action of these two thyroid-hormone-related compounds.     

Study of Ribavirin–Nucleic Acids Interaction

The UV–Vis absorption spectra of ribavirin in the absence and presence of calf thymus DNA are presented and discussed in this paper. The molecular structure of ribavirin was investigated by the semiempirical AM1 method, which triggered two polymorphic modifications of the antiviral drug also reported in the literature. Our experimental results point out two types of the binding. The first type involves a non-electrostatic (internal) binding, consisting of the intercalation of drug between the nitrogenous bases of nucleic acid. The second type (an external binding) involves the drug binding to the nucleic acid grooves. In addition, the binding constant of the second process has an order of magnitude greater than the binding constant for the first process, calculated by the Benesi–Hildebrand, Scott, and Scatchard methods, which supposes a 1:1 binding ratio. Also, the interactions of two polymorphic modifications of ribavirin (V₁ and V₂) with nucleic acids by the molecular mechanic and semiempirical AM1 methods were analyzed. The experimental data pointed out that in the ribavirin–nucleic acid complexes, the 1,2,4-triazole-3-carboxamide chromophore is intercalated between the bases of the nucleic acid sequences, the carboxamidic group is set outside of the nucleic acid sequence toward the major groove, and the 2-hydroxymethyl-tetrahydrofuran-3,4-diol fragment is located in the minor groove. In order to stress the sequence specificity of ribavirin, different models of the nucleic acid sequences containing adenine (A), thymine (T), cytosine (C), and guanine (G) in AAAAAA, TTTTTT, CCCCCC, GGGGGG, ATATAT, CGCGCG, ATCGAT, and CGATCG were used. The theoretical results outline the differences in the contributions of the electrostatic and van der Waals interactions to the total binding energy and the preference of ribavirin for the binding at the sequences of nucleic acids containing adenine and thymine bases.     

A comparative and mechanistic study on regioselective photooxidative demethylation of N‐heterocyclics using TiO2 and molecular oxygen

Regioselective photooxidative demethylation of methyl‐substituted N‐containing heterocyclic compounds is investigated in non‐aqueous (ethane nitrile) solutions containing semiconductor oxide (TiO₂) as photocatalyst in the presence of molecular oxygen upon irradiation at room temperature conditions. A plausible electron transfer mechanism, in which an electron‐hole pair is generated on the surface of TiO₂ by illumination, is proposed for the semiconductor‐mediated photocatalysis. Molecular orbital calculation by the AM 1 method has been performed for the electron densities of N‐atoms of heterocyclics in the present study. © 2002 Society of Chemical Industry     

Influence of Dicyclopentadiene Co-Polymers on the Hardening Processes and Properties of Portland Cement Composites

Dicyclopentadine (DCPD) polymers can be used for the manufacture of self healing materials due to their ability to form in the presence of a catalyst a highly cross-linked polycyclopendiene. The DCPD polymers selected for this work contain also another constituent sequence (maleic anhydride), therefore in this paper we assess also the possibility to use it for a potential self healing process. In this paper is presented the influence of two types of (DCPD) co-polymers based on maleic anhydride copolymers on the hardening processes and main properties of portland cement pastes and mortars, as a first step in the development of a self-healing cementitious composite. The properties (mechanical strengths and volume variations) of the cementitious composites with DCPD co-polymers are influenced by the chemical structure of the obtained compounds and curing medium (dry or humid air). The mortars with AM (maleic anhydride/dicyclopentadiene co-polymer) did not develop any mechanical strength due to an important swelling phenomenon determined by the hydrolysis of anhydride groups and sterical modifications in chain structures. The presence of APA (AM copolymer modified with p-amino phenyl acetic acid) decreases the compressive strengths but has a less important effect on the flexural strength. The APA polymer-cement interaction consists in a chelatization process involving the nitrogen atoms and hydrogen bonding of carboxyl groups providing, in this case, a good cohesion of the composite material. For both polymers a delay in cement hydration was noticed at early ages as well as the formation of organo-mineral phases.     

Design, syntheses, biological evaluation, and docking studies of 2-substituted 5-methylsulfonyl-1-phenyl-1H-indoles: potent and selective in vitro cyclooxygenase-2 inhibitors

Four series of 5-methylsulfonyl-1-phenyl-1H-indole-2-carboxylic acid alkyl esters (family A), -2-carbonitriles (family B), -2-carboxamides (family C), and 2-benzoyl-5-methylsulfonyl-1-phenyl-1H-indoles (family D) were prepared and evaluated for their ability to inhibit purified cyclooxygenase-2 (COX-2) and cyclooxygenase-1 (COX-1). Family D compounds have the best COX-1/COX-2 inhibition ratios and potencies. According to docking studies, these molecules appear to bind the COX-2 binding site differently than indomethacin, with the insertion of the substituent at the 2-position in the hydrophobic pocket of the enzyme and the 1-position phenyl ring in the trifluoromethyl zone. Among the group of compounds evaluated, 2-(4-chlorobenzoyl)-1-(4-chlorophenyl)-5-methylsulfonyl-1H-indole and 2-(4-chlorophenyl)-5-methylsulfonyl-1-(4-trifluoromethylphenyl)-1H-indole emerged as the most potent (respective IC(50) values: 46 and 43 nM), and selective (respective selectivity indexes: >2163 and >2331) COX-2 inhibitors.     

De Novo Design,Synthesis and Evaluation of Benzylpiperazine Derivatives as Highly Selective Binders of Mcl‐1

Considerable efforts have been made to the development of small‐molecule inhibitors of antiapoptotic B‐cell lymphoma 2 (Bcl‐2) family proteins (such as Bcl‐2, Bcl‐x_L, and Mcl‐1) as a new class of anticancer therapies. Unlike general inhibitors of the entire family, selective inhibitors of each member protein can hopefully reduce the adverse side effects in chemotherapy treatments of cancers overexpressing different Bcl‐2 family proteins. In this study, we designed four series of benzylpiperazine derivatives as plausible Bcl‐2 inhibitors based on the outcomes of a computational algorithm. A total of 81 compounds were synthesized, and their binding affinities to Bcl‐2, Bcl‐x_L, and Mcl‐1 measured. Encouragingly, 22 compounds exhibited binding affinities in the micromolar range (K_i<20 μM ) to at least one target protein. Moreover, some compounds were observed to be highly selective binders to Mcl‐1 with no detectable binding to Bcl‐2 or Bcl‐x_L, among which the most potent one has a K_i value of 0.18 μM for Mcl‐1. Binding modes of four selected compounds to Mcl‐1 and Bcl‐x_L were derived through molecular docking and molecular dynamics simulations. It seems that the binding affinity and selectivity of these compounds can be reasonably interpreted with these models. Our study demonstrated the possibility for obtaining selective Mcl‐1 inhibitors with relatively simple chemical scaffolds. The active compounds identified by us could be used as lead compounds for developing even more potent selective Mcl‐1 inhibitors with potential pharmaceutical applications.     

Synthesis and Evaluation of a Molecularly Imprinted Polymer for Solid Phase Extraction of Ethopabate from Chicken Tissue

In this paper the development and evaluation of a molecularly imprinted polymer (MIP) for ethopabate is described. Ethopabate (ETP), 4-acetamido-2-ethoxybenzoic acid methyl ester, is one of the antibiotics which is used as coccidiostat in poultry feeds. In the present study, two widely used functional monomers, methacrylic acid (MAA) and 4-vinylpyridine (4-VP) were compared theoretically and experimentally as the candidates for MIP preparation. Hyperchem software was employed to estimate binding energies between ETP and functional monomers and batch rebinding experiments were performed to study the binding characteristics of the polymers. The results showed that MAA is a better functional monomer to prepare MIP. UV/Vis and NMR spectroscopy were used as two common tools to study the interactions between ETP and MAA in the pre-polymerization mixture. Liquid chromatography experiments showed that the prepared MIP has recognition capability toward ETP in comparison with other structurally related compounds. The ETP-imprinted polymer was further applied for selective solid phase extraction (SPE) of ETP from a chicken tissue sample. The extraction yield of ETP was found to be quantitative (87 ± 3%) and the LOD and LOQ based on 3 and 10 times of the noise of HPLC profile were 0.05 and 0.32 ng ml^?1, respectively. It was confirmed that the binding ability of the prepared MIP for ETP was essentially sufficient in the presence of other compounds coexisting in tissue sample. Therefore, as a selective and efficient solid phase material, ETP-imprinted polymer has a high potential application in the analysis of residues of this antibiotic in chicken tissue samples.     

Towards Novel 3-Aminopyrazinamide-Based Prolyl-tRNA Synthetase Inhibitors: In Silico Modelling,Thermal Shift Assay and Structural Studies

Human cytosolic prolyl-tRNA synthetase (HcProRS) catalyses the formation of the prolyl-tRNA^Pro, playing an important role in protein synthesis. Inhibition of HcProRS activity has been shown to have potential benefits in the treatment of fibrosis, autoimmune diseases and cancer. Recently, potent pyrazinamide-based inhibitors were identified by a high-throughput screening (HTS) method, but no further elaboration was reported. The pyrazinamide core is a bioactive fragment found in numerous clinically validated drugs and has been subjected to various modifications. Therefore, we applied a virtual screening protocol to our in-house library of pyrazinamide-containing small molecules, searching for potential novel HcProRS inhibitors. We identified a series of 3-benzylaminopyrazine-2-carboxamide derivatives as positive hits. Five of them were confirmed by a thermal shift assay (TSA) with the best compounds 3b and 3c showing EC₅₀ values of 3.77 and 7.34 µM, respectively, in the presence of 1 mM of proline (Pro) and 3.45 µM enzyme concentration. Co-crystal structures of HcProRS in complex with these compounds and Pro confirmed the initial docking studies and show how the Pro facilitates binding of the ligands that compete with ATP substrate. Modelling 3b into other human class II aminoacyl-tRNA synthetases (aaRSs) indicated that the subtle differences in the ATP binding site of these enzymes likely contribute to its potential selective binding of HcProRS. Taken together, this study successfully identified novel HcProRS binders from our anti-tuberculosis in-house compound library, displaying opportunities for repurposing old drug candidates for new applications such as therapeutics in HcProRS-related diseases.     

Aurones as modulators of ABCG2 and ABCB1: synthesis and structure-activity relationships

The ability of aurones to modulate the efflux activities of ABCG2 and ABCB1 was investigated by quantifying their effects on the accumulation of pheophorbide?A (PhA) in ABCG2-overexpressing MDA-MB-231/R cells and calcein?AM in ABCB1-overexpressing MDCKII/MDR1 cells. Key structural features for interactions at both ABCG2 and ABCB1 are a methoxylated ring?A, an intact exocyclic double bond, and the location of the carbonyl bond on ring?C. Modifications on rings?B and C were less critical and served primarily to moderate activity and selectivity for one or both transporters. These SAR trends were quantified by Free-Wilson analyses and are reflected in a pharmacophore model for PhA accumulation. Several compounds were found to be equipotent with fumitremorgin?C (FTC) in promoting PhA accumulation, and they also demonstrated strong affinities for ABCB1. These compounds were disubstituted on ring?B with methoxy or a combination of methoxy and hydroxy groups. Taken together, our findings highlight the versatility of the aurone template as a lead scaffold for the design of dual-targeting ABCG2 and ABCB1 modulators.     

Fragments of gD Protein as Inhibitors of BTLA/HVEM Complex Formation - Design,Synthesis, and Cellular Studies

One of the major current trends in cancer immunotherapy is the blockade of immune checkpoint proteins that negatively regulate the immune response. This has been achieved through antibodies blocking PD-1/PD-L1 and CTLA-4/CD80/CD86 interactions. Such antibodies have revolutionized oncological therapy and shown a new way to fight cancer. Additional (negative) immune checkpoints are also promising targets in cancer therapy and there is a demand for inhibitors for these molecules. Our studies are focused on BTLA/HVEM complex, which inhibits T-cell proliferation and cytokine production and therefore has great potential as a new target for cancer treatment. The goal of the presented studies was the design and synthesis of compounds able to block BTLA/HVEM interactions. For that purpose, the N-terminal fragment of glycoprotein D (gD), which interacts with HVEM, was used. Based on the crystal structure of the gD/HVEM complex and MM/GBSA analysis performed on it, several peptides were designed and synthesized as potential inhibitors of the BTLA/HVEM interaction. Affinity tests, ELISA tests, and cellular-based reporter assays were performed on these compounds to check their ability to bind to HVEM and to inhibit BTLA/HVEM complex formation. For leading peptides candidates, all-atom and subsequent docking simulations with a coarse-grained force field were performed to determine their binding modes. To further evaluate their potential as drug candidates, their stability in plasma and their cytotoxicity effects on PBMCs were assessed. Our data indicate that the peptide gD(1-36)(K10C-T29C) is the best candidate as a future drug. It interacts with HVEM protein, blocks the BTLA/HVEM interaction, and is nontoxic to cells. The present study provides a new perspective on the development of BTLA/HVEM inhibitors that disrupt protein interactions.     

Selective agonists for dopamine/neurotensin receptor heterodimers

The neuromodulatory peptide neurotensin has been described to functionally interact with dopaminergic pathways of the human brain. We employed radioligand binding studies to investigate the physical interaction between co-expressed dopamine D(2L) or D? and neurotensin NTS? or NTS? receptors. Substantial cross-inhibitory effects of both receptor subtypes NTS(1) and NTS? on the agonist binding of D(2L) or D? were detected in the presence of neurotensin. To identify ligand-specific modulation and subtype-dependent differences, the novel dopamine receptor agonists 5 and 6 bearing the 7-OH-DPAT pharmacophore were synthesized. Exceptional ligand specificity was observed for D?-NTS? co-expression, which gave a 20-fold decrease in affinity for biphenylcarboxamide 5 in the presence of neurotensin. Comparing the binding properties of dopaminergic compounds in the presence of neurotensin, dopamine receptor subtype-selective profiles of the cross-inhibitory effect of neurotensin were observed.     

Influence of Complexation of Thiosemicarbazone Derivatives with Cu (II) Ions on Their Antitumor Activity against Melanoma Cells

A series of thiosemicarbazone derivatives was prepared and their anti-tumor activity in vitro was tested. The X-ray investigation performed for compounds T2, T3 and T5 confirmed the synthesis pathway and assumed molecular structures of analyzed thiosemicarbazones. The conformational preferences of the thiosemicarbazone system were characterized using theoretical calculations by AM1 method. Selected compounds were converted into complexes of Cu (II) ions. The effect of complexing on anti-tumor activity has been investigated. The copper(II) complexes, with Schiff bases T1, T10, T12, T13, and T16 have been synthesized and characterized by chemical and elemental analysis, FTIR spectroscopy and TGA method. Thermal properties of coordination compounds were studied using TG-DTG techniques under dry air atmosphere. G361, A375, and SK-MEL-28 human melanoma cells and BJ human normal fibroblast cells were treated with tested compounds and their cytotoxicity was evaluated with MTT test. The compounds with the most promising anti-tumour activity were then selected and their cytotoxicity was verified with cell cycle analysis and apoptosis/necrosis detection. Additionally, DNA damages in the form of a basic sites presence and the expression of oxidative stress and DNA damage response genes were evaluated. The obtained results indicate that complexation of thiosemicarbazone derivatives with Cu (II) ions improves their antitumor activity against melanoma cells. The observed cytotoxic effect is associated with DNA damage and G2/M phase of cell cycle arrest as well as disorders of the antioxidant enzymes expression.     

Cationic activated monomer polymerization of heterocyclic monomers

In the first part of this review the meaning of activation is discussed and selected examples of polymerizaton processes in which activation of monomer is required prior to actual propagation are presented. In some systems, activation of monomer proceeds with such a strong interaction between an activator and monomer that a new chemical entity is derived from the monomer. To describe the mechanism of such a process, the term ‘Activated Monomer Mechanism’ has been coined.

The main part of the review is concerned with cationic Activated Monomer (AM) polymerization of cyclic ethers. In this process, cyclic ether is activated by formation of protonated species in the presence of a protic acid. Reaction of the protonated (activated) cyclic ether with hydroxyl group containing compounds leads to ring opening reforming the hydroxyl group. Several repetitions of such a reaction constitute a chain process. Thus, in AM polymerization of cyclic ethers hydroxyl group containing compounds act as initiator, protic acid is a catalyst, growing chain end is fitted with hydroxyl group and the charged species is a protonated monomer. The important feature of such a polymerization mechanism is that due to the absence of charged species at the growing chain end, back-biting leading to the formation of macrocyclics can be eliminated.

The mechanism and kinetics of AM polymerization of cyclic ethers is discussed and the approach allowing one to determine the rate constant for propagation involving activated monomer species is outlined. The application of the AM concept to the copolymerization of cyclic ethers as well as to the polymerization of monomers containing both initiating (hydroxyl groups) and propagating (cyclic ether) functions within one molecule are presented.

In the subsequent parts of the review, examples of cationic AM polymerization of other types of heterocyclic monomers, including cyclic acetals, cyclic esters (lactones), amines and amides (lactams), are given.

Finally, the polyaddition of oxiranes to derivatives of phosphoric acid is discussed. Although this system does not conform to the AM polymerization scheme, it bears formal resemblance to earlier systems in such a sense that the activation of the cyclic ether is required for the reaction to occur.     

Water-soluble copolymers: 22. Copolymers of acrylamide with 2-acrylamido-2-methylpropanedimethylammonium chloride: Aqueous solution properties of a polycation

The viscometric, turbidimetric and potentiometric properties of copolymers of acrylamide (AM) with 2-acrylamido-2-methylpropanedimethylammonium chloride (AMPDAC) were studied in aqueous solutions. The AMPDAC polymers exhibit poor salt tolerance and large, negative viscosity/temperature coefficients. Furthermore, the polymers were found to be sensitive to changes in pH. The AMPDAC polymers undergo phase separation in the presence of dianions as a function of temperature and AMPDAC composition.     

Molecular imprinted solid‐phase extraction of huperzine A from Huperzia Serrata

On the basis of the non‐covalent interaction between template and monomer, porous molecularly imprinted polymers (MIPs) were synthesized by a thermal‐initiated polymerization method using huperzine A as template, acrylamide, or methacrylic acid as function monomer, ethylene glycol dimethacrylate as cross‐linking agent. The interaction between template and functional monomers was studied by UV spectrophotometry, which showed a formation of huperzine A‐monomer complexes with stoichiometric ratio of 1 : 2 in the pre‐polymerized systems. The resultant MIP particles were tested in the equilibrium binding experiment to analyze their adsorption ability to huperzine A, and were characterized by Fourier Transform Infrared (FTIR) study. The recognition properties of MIP were estimated in solid‐phase extraction by selecting four compounds (isolated from the Chinese herb Huperzia serrata) as substrates, and were compared with and prior to those of the NIP. High affinity and adsorption of MIP1 which was prepared in chloroform with huperzine A as imprinted molecule, and acrylamide (AM) as functional monomer, made an attractive application of MIP1 in separation processes. In final, using MIP1 solid‐phase extraction micro‐column, huperzine A was enriched and separated from the real extraction sample of Huperzia serrata. © 2009 Wiley Periodicals, Inc. J Appl Polym Sci, 2009