Epoxide Species Research Articles

Halogenation of Alkenes Using Three‐Component Reactions: A Decade of Development

AbstractAlkenes are valuable feedstocks in organic synthesis. One effective method for synthesizing organic halides with functional groups in close proximity involves the direct difunctionalization of alkenes via three‐component reactions. This approach not only reduces the number of steps involved in the synthesis process, but also minimizes waste generation and enables the formation of complex molecules from simple starting materials. In this review, we mainly discuss decade developments (2013‐2023) in two categories: (1) halogenation via three‐membered ring intermediates, involving haliranium, thiiranium,seleniranium, aziridinium and epoxide species; (2) halogenation via a radical pathway. Reactions with I2, BiI3, NaI, Bu4N+[I(O2CAr)2]−, NIS, NBS, NCS, DBH, BsNMeBr, HBr, HCl, KI, NH4I, I2O5, Et3N ⋅ 3HF, Selectfluor, CuI, CuBr, CuCl, LiCl, KBr, NaCl, SOCl2, Py ⋅ 9HF, NFSI, TBSCl et al have been recorded and how the added reagents work will be discussed. We hope this review will do help for future research in this area.

Serum Arachidonic Acid Oxylipins Indicate Small Vessel Injury And the Related Executive Function Decline In Stroke Patients

AbstractBackgroundCerebral small vessel disease (SVD) frequently co‐occurs with other neurodegenerative pathologies and collectively lead to cognitive deficits. Previously, we found that the ratios of soluble epoxide hydrolase (sEH)‐ derived diols to the parent epoxides (generated by CYP2J/2Cs) linoleic acids were associated with greater white matter hyperintensities (WMH) and poorer executive function in patients with transient ischemic attack. However, the associations with the diol and epoxide species derived from arachidonic acid (AA) have not been explored.MethodsThe study cohort is comprised of 159 stroke patients (mean age = 69.2, 32.1% female) from the Ontario Neurodegenerative Degenerative Research Initiative. EpETrEs (AA epoxides) and DiHETrEs (AA diols) were quantified by a targeted ultrahigh pressure liquid chromatography mass spectrometry (UPLC‐MS/MS). Executive function was assessed using a composite score that included Digit Symbol Substitution Test (DSST), Trial‐Making Test Part B, Stroop Color‐Word Interference Test, and FAS Verbal Fluency Test. WMHs were quantified from T2‐Fluid‐Attenuated‐Inversion‐Recovery MRI through Lesion Explorer (https://imaging.brainlab.ca/lesion‐explorer.html).In linear models controlling for age, sex, education or age, sex, glucose, cholesterol, and waist circumferences were used for cognitive and imaging out comes, respectively.Results11(12)‐EpETrE was found to be associated with less WHM (ρ = ‐0.175, p = 0.040) and better executive function (ρ = 0.166, p = 0.039). In a sub‐cohort of patients with small vessel stroke (n = 50), the relationship between WMH and 11(12)‐EpETrE became stronger (ρ = ‐0.390, p = 0.014). 14(15)‐EpETrE was also associated with less WMH (ρ = ‐0.416, p = 0.009). Similar relationships were not seen with the diols or diol to epoxide ratios.ConclusionsOxylipins derived from sEH activity may be potential biomarkers of vascular cognitive impairment and small vessel disease in stroke patients.

Metabolite V, an epoxide species is a minor circulating metabolite in humans following a single oral dose of deflazacort.

Deflazacort (Emflaza) was approved in the United States in 2017 for the treatment of the Duchenne muscular dystrophy in patients aged 2 years and older. Several deflazacort metabolites were isolated and identified from rats, dogs, monkeys, and humans. Among them, 1ß,2ß‐epoxy‐3ß‐hydroxy‐21‐desacetyl deflazacort, referred to as Metabolite V, was reported to be one of the major circulating metabolites in humans. However, its quantitative distribution in plasma was not fully characterized. The objective of this study was to determine deflazacort plasma pharmacokinetics, metabolite profiles and their quantitative exposures in humans following a single oral dose. Six healthy male subjects were each administered a single oral dose of 60 mg [14C]‐deflazacort. Plasma and urine were collected and deflazacort metabolites in plasma were quantified by high performance liquid chromatography radio‐profiling followed by liquid chromatography‐mass spectrometry characterization. Metabolite V was isolated from urine and its structure was further confirmed by nuclear magnetic resonance analysis. These analyses demonstrated that deflazacort was not detectable in plasma; of the eight circulating deflazacort metabolites identified or characterized, the pharmacologically active metabolite 21‐desacetyl deflazacort and inactive metabolite 6ß‐hydroxy‐21‐desacetyl deflazacort accounted for 25.0% and 32.9% of the 0‐24 hours plasma total radioactivity, respectively, while Metabolite V, an epoxide species, was a minor circulating metabolite, representing only about 4.7% of the total plasma radioactivity.

A novel concept for the biodegradation mechanism of dianionic catechol with homoprotocatechuate 2,3-dioxygenase: A non-proton-assisted process.

The theory of "proton-assisted process" can well explain the catalytic mechanism of homoprotocatechuate 2,3-dioxygenase (2,3-HPCD) with a monoanionic substrate (homoprotocatechuate, HPCA). Here a "non-proton-assisted process" is presented to interpret catalytic mechanism of 2,3-HPCD with a dianionic substrate (4-nitrocatechol, 4NC). The ONIOM calculation is performed to investigate the reaction pathway of a wild-type 2,3-HPCD with 4NC (H200H-4NC system). The catalytic reaction is comprised of four steps: (1) A dioxygen attacks the aromatic ring to produce an alkylperoxo species. (2) O-O bond cleavage and the formation of an epoxide species occur. (3) A seven-membered O-heterocyclic compound is generated by the extinction of the epoxy structure. (4) The seven-membered ring undergoes ring opening to form the final product (C2-C3 cleavage product). The effective free energy barrier of the catalytic reaction of the H200H-4NC system is 26.2kcalmol-1, which is much higher than that of the H200H-HPCA system. Furthermore, two calculated electronic configurations (Fe(III)-O2•- and Fe(III)-SQ•) have a high similarity to previously detected ones, which demonstrates that the Asn200 variant (H200N-4NC variant system) employs a C4 (para-carbon) pathway to produce a C4-C5 cleavage product. Our findings provide an in-depth understanding of the catalytic mechanisms of dianionic catechol and its derivatives.

Identification of Structural‐Activity Features of Long Chain Polyunsaturated Fatty Acid Epoxides in Angiogenesis

The cytochrome P450‐derived fatty acid epoxides are bioactive lipids with regulatory roles in a wide range of homeostatic and pathophysiological events. These lipid mediators are metabolized efficiently by the mammalian soluble epoxide hydrolase enzyme (sEH) to corresponding 1,2‐diols. Inhibition of sEH has been shown to be beneficial to human health via increasing concentrations of a variety of short‐lived epoxylipids in vivo (2). Identifying the most potent polyunsaturated fatty acid (PUFA) epoxide metabolites will provide insight on the effects of sEH inhibition. Recent studies have shown that the omega‐6 epoxyeicosatrienoic acids (EETs) from arachidonic acid are pro‐angiogenic, while the omega‐3, docosahexaenoic acid (DHA)‐derived epoxydocosapentaenoic acids (EDPs) inhibit angiogenesis, tumor growth, and metastasis (1). Given these findings, we are interested in exploring the roles of various long chain PUFA epoxides in angiogenesis. We tested a library of omega‐3 and omega‐6 epoxides in a newly developed high‐throughput tube formation assay and found their effects are regiospecific and stereospecific; Of note, the 19(S),20(R)‐EDP stereoisomer was a less effective inhibitor of endothelial cell tube formation than its respective racemic mixture. Here, we report the angiogenic activities of diverse omega‐3 and omega‐6 fatty acid epoxides that could be present in mammals from the diet, including the less explored epoxides from both omega‐3 and omega‐6 docosapentaenoic acid (DPA). We will discuss the structure‐activity features of the most potent epoxide species and also present methodology to synthesize and purify the regioisomers and stereoisomers of each series of epoxides. Ultimately, understanding activities of sEH substrates in specific regulatory environments will guide development of targeted sEH modulators that promote the beneficial effects of PUFA metabolites in human health and disease.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Facile and Direct Hydroxylation of Benzene to Phenol over Graphene‐Based Catalysts: Integrated Utilization of Greenhouse Nitrous Oxide

AbstractGraphene catalysis has recently emerged as a focus and herein a novel and highly efficient process is computationally designed using graphene‐based catalysts that integrates the utilization of greenhouse nitrous oxide (N2O) as oxidant. The ground‐state pristine graphene is not catalytically active for benzene hydroxylation, and doping and photocatalysis through excitations are employed to improve the catalyst systems, while both strategies exhibit limited effects. A combination of Be‐doping and photocatalysis through excitations results in encouraging changes, where all energy barriers are moderate, so that the whole catalytic processes proceed facilely at mild conditions. Graphene materials, especially Be‐ and O‐doped ones, are good photocatalysts due to the low excitation energies. Mechanistic studies indicate that the active sites of the excited‐state graphene, whether pristine or doped, are oxygen anion radical (O−•) and distinct from the epoxide species for the ground state; in addition, Be‐doping pronouncedly reduces the excitation energies and enhances the interactions with the active‐site O species, which further show strong stabilization effects to transition states and reduce substantially the energy barriers.

Study of the Synchrotron Photoionization Oxidation of 2-Methylfuran Initiated by O(3P) under Low-Temperature Conditions at 550 and 650 K.

The O-(3P)-initiated oxidation of 2-methylfuran (2-MF) was investigated using vacuum-ultraviolet synchrotron radiation from the Advanced Light Source at Lawrence Berkeley National Laboratory. Reaction species were studied by multiplexed photoionization mass spectrometry at 550 and 650 K. The oxygen addition pathway is favored in this reaction, forming four triplet diradicals that undergo intersystem crossing into singlet epoxide species that lead to the formation of products at m/z 30 (formaldehyde), 42 (propene), 54 (1-butyne, 1,3-butadiene, and 2-butyne), and 70 (2-butenal, methyl vinyl ketone, and 3-butenal). Mass-to-charge ratios, photoionization spectra, and adiabatic ionization energies for each primary reaction species were obtained and used to characterize their identities. In addition, by means of electronic structure calculations, potential energy surface scans of the different species produced throughout the oxidation were examined to further validate the primary chemistry occurring. Branching fractions for the formation of the primary products were calculated at the two temperatures and contribute 81.0 ± 21.4% at 550 K and 92.1 ± 25.5% at 650 K.

Amine-free melanin-concentrating hormone receptor 1 antagonists: Novel 1-(1H-benzimidazol-6-yl)pyridin-2(1H)-one derivatives and design to avoid CYP3A4 time-dependent inhibition

Melanin-concentrating hormone (MCH) is an attractive target for antiobesity agents, and numerous drug discovery programs are dedicated to finding small-molecule MCH receptor 1 (MCHR1) antagonists. We recently reported novel pyridine-2(1H)-ones as aliphatic amine-free MCHR1 antagonists that structurally featured an imidazo[1,2-a]pyridine-based bicyclic motif. To investigate imidazopyridine variants with lower basicity and less potential to inhibit cytochrome P450 3A4 (CYP3A4), we designed pyridine-2(1H)-ones bearing various less basic bicyclic motifs. Among these, a lead compound 6a bearing a 1H-benzimidazole motif showed comparable binding affinity to MCHR1 to the corresponding imidazopyridine derivative 1. Optimization of 6a afforded a series of potent thiophene derivatives (6q–u); however, most of these were found to cause time-dependent inhibition (TDI) of CYP3A4. As bioactivation of thiophenes to form sulfoxide or epoxide species was considered to be a major cause of CYP3A4 TDI, we introduced electron withdrawing groups on the thiophene and found that a CF3 group on the ring or a Cl adjacent to the sulfur atom helped prevent CYP3A4 TDI. Consequently, 4-[(5-chlorothiophen-2-yl)methoxy]-1-(2-cyclopropyl-1-methyl-1H-benzimidazol-6-yl)pyridin-2(1H)-one (6s) was identified as a potent MCHR1 antagonist without the risk of CYP3A4 TDI, which exhibited a promising safety profile including low CYP3A4 inhibition and exerted significant antiobesity effects in diet-induced obese F344 rats.

Atomic Layer Deposition of Hafnium(IV) Oxide on Graphene Oxide: Probing Interfacial Chemistry and Nucleation by using X-ray Absorption and Photoelectron Spectroscopies.

Interfacing graphene with metal oxides is of considerable technological importance for modulating carrier density through electrostatic gating as well as for the design of earth-abundant electrocatalysts. Herein, we probe the early stages of the atomic layer deposition (ALD) of HfO2 on graphene oxide using a combination of C and O K-edge near-edge X-ray absorption fine structure spectroscopies and X-ray photoelectron spectroscopy. Dosing with water is observed to promote defunctionalization of graphene oxide as a result of the reaction between water and hydroxyl/epoxide species, which yields carbonyl groups that further react with migratory epoxide species to release CO2 . The carboxylates formed by the reaction of carbonyl and epoxide species facilitate binding of Hf precursors to graphene oxide surfaces. The ALD process is accompanied by recovery of the π-conjugated framework of graphene. The delineation of binding modes provides a means to rationally assemble 2D heterostructures.

Gold-decorated graphene nanosheets composed of a biocompatible non-charged water-soluble polypeptide

Nanocomposite materials derived from carbon/polymer hybrids and metallic nanoparticles are of great interest due to the wide spectrum of potential applications. In this work, the biocompatible, non-charged and independent of pH, water soluble polypeptide, poly(L-proline) (PLP), was covalently grafted on exfoliated graphene oxide (GO) by a ‘grafting to’ approach, through the ring opening reaction of the surface epoxide species, enabling thus the simultaneous reduction and stabilization of as-synthesized gold nanoparticles in water in one-step, without any toxic by-product formation. The PLP-mediated GO nanocomposites serve as a new scaffold for metallic particles supported on a carbon matrix through the polypeptide mediation, and could give rise to multiple applications, in catalysis and cancer therapies.

Radical-Based Epoxide Opening by Titanocenes

The binding of 2,2-diphenyloxirane to Cp2TiCl is studied on the electronic level by magnetic resonance spectroscopy and quantum chemical calculations. The complexation of 2,2-diphenyloxirane is accompanied by dissociation of the chloride ligand, and thus, the epoxide binds to the cationic titanocene(III) complex. The titanocene(III)-epoxide species persists only for short periods of time (<5 min) even at 243 K, indicating that the ring-opening reaction is exothermic. A short-lived paramagnetic titanocene(IV)-epoxide radical species has not been directly observed. However, by a combination of isotope labeling and spin-trapping, evidence for the existence of such a species has been unequivocally demonstrated. The observation of a titanocene(III)-epoxide complex is unprecedented and provides direct evidence for inner-sphere electron transfer between epoxides and titanocenes, responsible for the high regioselectivity of ring-opening.

Hybrid peroxotungstophosphate organized catalysts highly active and selective in alkene epoxidation

Heterogenization of homogeneous catalysts is still a challenge but can improve drastically the processability of these compounds. Hybridization of polyoxometalates offers an efficient heterogenization route of homogeneous epoxidation catalysts. The Keggin [PW12O40]3 − and the Ventruello {PO4[WO(O2)2]4}3 − species were inserted by electrostatic interactions in a poly(ampholytic) polymeric matrix in order to prevent the leaching of these species. The presence of the polymeric matrix allows to tune the catalyst performance in cyclooctene epoxidation and to improve the selectivity to epoxide. Indeed, the hydrophobicity of the matrix induces a quick desorption of hydrophilic epoxide species. Their over-oxidation and the catalyst species deactivation by over-adsorption of epoxide are then avoided.

A novel hydrofobic niobium oxyhydroxide as catalyst: Selective cyclohexene oxidation to epoxide

A surface modified niobium oxyhydroxide catalyst was utilized for the selective liquid-phase oxidation of cyclohexene to epoxide in the presence of hydrogen peroxide. After surface modification with a surfactant (cetiltrimetilamonium bromide -CTAB), the very active niobium oxyhydroxide (NbO2OH), presented hydrophobic characteristics and good activity for the oxidation of cyclohexene over the heterogeneous catalyst. Furthermore, the hydrophobic material presented a high selectivity toward the formation of the epoxide species. The mass spectrometry analysis of the reaction showed that epoxide species was produced with a selectivity of 60% using 0.2mL of hydrogen peroxide, with 65% of conversion. The results strongly suggest that the reaction involves oxidizing species generated after the reaction with H2O2. The surfactant anchored over the niobium catalyst promotes a better interaction with the nonpolar substrate.

HO2 formation from the OH + benzene reaction in the presence of O2

In this study we investigated the secondary formation of HO(2) following the benzene + OH reaction in N(2) with variable O(2) content at atmospheric pressure and room temperature in the absence of NO. After pulsed formation of OH, HO(x) (= OH + HO(2)) and OH decay curves were measured by means of a laser-induced fluorescence technique (LIF). In synthetic air the total HO(2) yield was determined to be 0.69 ± 0.10 by comparison to results obtained with CO as a reference compound. HO(2) is expected to be a direct product of the reaction of the intermediately formed OH-benzene adduct with O(2). The HO(2) yield is slightly greater than the currently recommended yield of the proposed HO(2) co-product phenol (∼53%). This hints towards other, minor HO(2) forming channels in the absence of NO, e.g. the formation of epoxide species that was proposed in the literature. For other test compounds upper limits of HO(2) yields of 0.10 (isoprene) and 0.05 (cyclohexane) were obtained, respectively. In further experiments at low O(2) concentrations (0.06-0.14% in N(2)) rate constants of (2.4 ± 1.1) × 10(-16) cm(3) s(-1) and (5.6 ± 1.1) × 10(-12) cm(3) s(-1) were estimated for the OH-benzene adduct reactions with O(2) and O(3), respectively. The rate constant of the unimolecular dissociation of the adduct back to benzene + OH was determined to be (3.9 ± 1.3) s(-1). The HO(2) yield at low O(2) was similar to that found in synthetic air, independent of O(2) and O(3) concentrations indicating comparable HO(2) yields for the adduct + O(2) and adduct + O(3) reactions.

On the observation of a gem diol intermediate after O–O bond cleavage by extradiol dioxygenases. A hybrid DFT study

Catalytic cycle intermediates of a representative extradiol dioxygenase, homoprotocatechuate 2,3-dioxygenase (HPCD), have recently been characterized in crystallo by Kovaleva and Lipscomb. The structures of the identified species indicate that the process of inserting oxygen into the catechol ring occurs stepwise, and involves an Fe(II)-alkylperoxo intermediate and its O-O cleavage product: a gem diol species. In general, these findings corroborate the results of our previous computational studies; however, the fact that the gem diol species is stable enough to be observed in the crystal form seems to be at odds with the computational mechanistic data, which suggest that this intermediate should very readily and spontaneously convert to the epoxide species. The key question then becomes what is actually observed in the X-ray experiments. Here we report additional computational studies undertaken with the hope of clarifying this issue. The results obtained for active site models hosting both the native and the alternative (4-sulfonylcatechol) substrate indicate that the stability of the gem diol species is substantially increased if an electron and a proton are added. If this occurs somehow, the lifetime of the intermediate should be sufficient to observe it.

Selection of high-affinity human monoclonal antibodies specific to the constant domain of versican as tools for tumor targeting

Melanin-concentrating hormone (MCH) is an attractive target for antiobesity agents, and numerous drug discovery programs are dedicated to finding small-molecule MCH receptor 1 (MCHR1) antagonists. We recently reported novel pyridine-2(1H)-ones as aliphatic amine-free MCHR1 antagonists that structurally featured an imidazo[1,2-a]pyridine-based bicyclic motif. To investigate imidazopyridine variants with lower basicity and less potential to inhibit cytochrome P450 3A4 (CYP3A4), we designed pyridine-2(1H)-ones bearing various less basic bicyclic motifs. Among these, a lead compound 6a bearing a 1H-benzimidazole motif showed comparable binding affinity to MCHR1 to the corresponding imidazopyridine derivative 1. Optimization of 6a afforded a series of potent thiophene derivatives (6q–u); however, most of these were found to cause time-dependent inhibition (TDI) of CYP3A4. As bioactivation of thiophenes to form sulfoxide or epoxide species was considered to be a major cause of CYP3A4 TDI, we introduced electron withdrawing groups on the thiophene and found that a CF3 group on the ring or a Cl adjacent to the sulfur atom helped prevent CYP3A4 TDI. Consequently, 4-[(5-chlorothiophen-2-yl)methoxy]-1-(2-cyclopropyl-1-methyl-1H-benzimidazol-6-yl)pyridin-2(1H)-one (6s) was identified as a potent MCHR1 antagonist without the risk of CYP3A4 TDI, which exhibited a promising safety profile including low CYP3A4 inhibition and exerted significant antiobesity effects in diet-induced obese F344 rats.

Toward a myeloablative regimen with clinical potential: II. Treosulfan induces specific skin graft tolerance across haploidentical MHC barriers.

Treosulfan is a water-soluble structural analog of busulfan, acting as a prodrug of alkylating epoxide species. It does not induce severe hepatotoxicity or veno-occlusive disease at or above the maximum tolerated dose, lacks significant nonhematological toxicity and has a limited organ toxicity. It is mainly indicated for the treatment of patients with ovarian cancer. In the present study, we report that permanent donor-specific tolerance and stable mixed multilineage chimerism can successfully be achieved across haploidentical MHC barriers when Treosulfan is administered in combination with anti-T-cell mAb and T-cell-depleted donor bone marrow cells. Furthermore, we show that less T-cell suppression is required when Treosulfan is included in the conditioning regimen. In conclusion, Treosulfan is a well-tolerated myeloablative agent with a low toxicity, and is a promising candidate drug for conditioning prior to bone marrow transplantation.

Tissue-Specific Utilization of Menaquinone-4 Results in the Prevention of Arterial Calcification in Warfarin-Treated Rats

The effects of vitamin K (phylloquinone: K1 and menaquinone-4: MK-4) on vascular calcification and their utilization in the arterial vessel wall were compared in the warfarin-treated rat model for arterial calcification. Warfarin-treated rats were fed diets containing K1, MK-4, or both. Both K1 and MK-4 are cofactors for the endoplasmic reticulum enzyme γ-glutamyl carboxylase but have a structurally different aliphatic side chain. Despite their similar in vitro cofactor activity we show that MK-4 and not K1 inhibits warfarin-induced arterial calcification. The total hepatic K1 accumulation was threefold higher than that of MK-4, whereas aortic MK-4 was three times that of K1. The utilization of K1 and MK-4 in various tissues was estimated by calculating the ratios between accumulated quinone and epoxide species. K1 and MK-4 were both equally utilized in the liver, but the aorta showed a more efficient utilization of MK-4. Therefore, the observed differences between K1 and MK-4 with respect to inhibition of arterial calcification may be explained by both differences in their tissue bioavailability and cofactor utilization in the reductase/carboxylase reaction. An alternative explanation may come from an as yet hypothetical function of the geranylgeranyl side chain of MK-4, which is a structural analogue of geranylgeranyl pyrophosphate and could interfere with a critical step in the mevalonate pathway.

Amphiphilic copolymers of glycidol with nonpolar epoxide comonomers

AbstractTo explore the industrially used copolymerization of glycidol with other epoxide species, a series of copolymers of glycidol with various comonomers (epichlorohydrin, isopropyl glycidyl ether, 1,2‐epoxybutane, propylene oxide, and glycidyl phenyl ether) has been synthesized by cationic ring‐opening polymerization in dichloromethane, using boron trifluoride diethyl etherate as the initiator. The facile synthesis proceeded at room temperature under ordinary atmosphere. The air‐ and temperature‐stable products—mostly clear, colorless, viscous liquids—are proposed to consist of a hyperbranched polyglycidol core (incorporating varying fractions of comonomer) with arms made from comonomer‐derived repeat units. These copolymers had low molecular weights and rather broad molecular weight distributions. There was wide variation in the polarity of these copolymers, depending on the comonomer used, resulting in materials that were soluble in solvents ranging from benzene to water. The glass‐transition temperatures observed for these copolymers also varied widely, ranging from −56 to 1°C. © 2001 John Wiley &amp; Sons, Inc. J Appl Polym Sci 82: 2290–2299, 2001

DNA alkylation and interstrand cross-linking by treosulfan.

The anti-tumour drug treosulfan (L-threitol 1,4-bismethanesulphonate, Ovastat) is a prodrug for epoxy compounds by converting non-enzymatically to L-diepoxybutane via the corresponding monoepoxide under physiological conditions. The present study supports the hypothesis that this conversion of treosulfan is required for cytotoxicity in vitro. DNA alkylation and interstrand cross-linking of plasmid DNA is observed after treosulfan treatment, but this is again produced via the epoxide species. Alkylation occurs at guanine bases with a sequence selectivity similar to other alkylating agents such as the nitrogen mustards. In treosulfan-treated K562 cells, cross-links form slowly, reaching a peak at approximately 24 h. Incubation of K562 cells with preformed epoxides shows faster and more efficient DNA cross-linking. © 1999 Cancer Research Campaign