Hydrolysis Of Lactams Research Articles

Biodegradation of penicillin G sodium by Sphingobacterium sp. SQW1: Performance, degradation mechanism, and key enzymes

Biodegradation is an efficient and cost–effective approach to remove residual penicillin G sodium (PGNa) from the environment. In this study, the effective PGNa–degrading strain SQW1 (Sphingobacterium sp.) was screened from contaminated soil using enrichment technique. The effects of critical operational parameters on PGNa degradation by strain SQW1 were systematically investigated, and these parameters were optimized by response surface methodology to maximize PGNa degradation. Comparative experiments found the extracellular enzyme to completely degrade PGNa within 60 min. Combined with whole genome sequencing of strain SQW1 and LC–MS analysis of degradation products, penicillin acylase and β–lactamase were identified as critical enzymes for PGNa biodegradation. Moreover, three degradation pathways were postulated, including β–lactam hydrolysis, penicillin acylase hydrolysis, decarboxylation, desulfurization, demethylation, oxidative dehydrogenation, hydroxyl reduction, and demethylation reactions. The toxicity of PGNa biodegradation intermediates was assessed using paper diffusion method, ECOSAR, and TEST software, which showed that the biodegradation products had low toxicity. This study is the first to describe PGNa–degrading bacteria and detailed degradation mechanisms, which will provide new insights into the PGNa biodegradation.

An efficient, concise synthetic approach to γ-aminobutyric acid (GABA) derivatives bearing bicyclo[2.2.2]octane and bicyclo[2.2.1]heptane rings

An efficient, concise 5-step synthetic approach to γ-aminobutyric acid (GABA) derivatives bearing bridged bicyclo[2.2.2]octane and bicyclo[2.2.1]heptane rings was developed, which consists of Diels-Alder cycloaddition to construct the bridged bicyclic rings with a nitrile functionality, hydrogenation, LDA-mediated SN2 alkylation to introduce acetate moiety at the α-position of nitrile, reduction of the nitrile functionality to amino group followed by intramolecular formation of lactam and finally acidic hydrolysis of lactam to give the desired GABA derivatives (1 and 19 as HBr salts). The reaction conditions of Diels-Alder cycloaddition and LDA-mediated SN2 alkylation were intensively optimized to improve the yields. The stereochemistry of the SN2 reaction involved in the bicyclo[2.2.1]heptane ring system was unambiguously elucidated by single-crystal X-ray diffraction. This synthetic approach possesses advantages of less reaction steps, high overall yields and avoidance of toxic reagents, and may find wide applications in the synthesis of other GABA derivatives with similar bicyclic or polycyclic rings.

Novel amino acid ionic liquids prepared via one‐step lactam hydrolysis for the highly efficient capture of CO2

AbstractThis article proposes for the first time the preparation of a series of amino acid ionic liquids (AAILs) via one‐step hydrolysis of cheap lactams for the capture of CO2. The structures of the prepared AAILs are confirmed using NMR, FTIR, and ESI‐MS, and their physical properties are also determined. It is found that these AAILs are reversible CO2 absorbents with very high absorption capacities (0.15 to 0.18 g·g−1 at 313.2 K and 1.0 bar), better than almost all task‐specific ionic liquids reported in literatures. The absorption mechanism is also elucidated to be a combination of 1:1 and 2:1 stoichiometric reaction of AAILs with CO2 from NMR, FTIR, reaction equilibrium thermodynamical modeling and quantum calculations. The AAILs have the advantages of simple synthesis, high yield, and using available cheap raw materials. It is believed that this kind of AAILs have great potentials to be used as efficient CO2 absorbents.

Mass Spectrometry-Based Detection of Beta Lactam Hydrolysis Enables Rapid Detection of Beta Lactamase Mediated Antibiotic Resistance.

Antibiotic resistance by beta lactamase expression is a serious and growing threat. We aimed to determine whether beta-lactamase activity is detectable in urine specimens to enable faster identification of resistance. Urine specimens from patients with extended spectrum beta lactamase (ESBL)-expressing urinary infections were incubated with beta lactam antibiotics. Beta lactam hydrolysis was determined by mass spectrometry methods. Ceftriaxone hydrolysis was observed in 45 of 45 ESBL-containing specimens from patients not treated with a beta lactamase inhibitor before specimen collection. Ceftriaxone hydrolysis was not observed in 108 of 108 non-ESBL-containing specimens. Spiking studies show that beta lactam hydrolysis can be observed within 30 minutes. Beta lactam hydrolysis is evidenced by mass spectrometry preceded by either liquid chromatography or matrix-assisted laser desorption ionization specimen processing methods. Clinically significant beta lactamase activity is detectable directly from urine specimens. The described methods would enable the detection of beta lactam resistance 24 to 48 hours sooner than culture based methods.

Catalytic and structural properties of ATP-dependent caprolactamase from Pseudomonas jessenii.

Caprolactamase is the first enzyme in the caprolactam degradation pathway of Pseudomonas jessenii. It is composed of two subunits (CapA and CapB) and sequence‐related to other ATP‐dependent enzymes involved in lactam hydrolysis, like 5‐oxoprolinases and hydantoinases. Low sequence similarity also exists with ATP‐dependent acetone‐ and acetophenone carboxylases. The caprolactamase was produced in Escherichia coli, isolated by His‐tag affinity chromatography, and subjected to functional and structural studies. Activity toward caprolactam required ATP and was dependent on the presence of bicarbonate in the assay buffer. The hydrolysis product was identified as 6‐aminocaproic acid. Quantum mechanical modeling indicated that the hydrolysis of caprolactam was highly disfavored (ΔG0'= 23 kJ/mol), which explained the ATP dependence. A crystal structure showed that the enzyme exists as an (αβ)2 tetramer and revealed an ATP‐binding site in CapA and a Zn‐coordinating site in CapB. Mutations in the ATP‐binding site of CapA (D11A and D295A) significantly reduced product formation. Mutants with substitutions in the metal binding site of CapB (D41A, H99A, D101A, and H124A) were inactive and less thermostable than the wild‐type enzyme. These residues proved to be essential for activity and on basis of the experimental findings we propose possible mechanisms for ATP‐dependent lactam hydrolysis.

Enantioselective Total Synthesis of Desbromoarborescidines A–C and the Formal Synthesis of (S)-Deplancheine

Starting from Boc-protected tryptamine and (S)-tetrahydro-5-oxo-2-furancarboxylic acid, facile enantioselective total synthesis of desbromoarborescidines A-C and the formal synthesis of (S)-deplancheine have been accomplished via a common intermediate (S)-indolo[2,3-a]quinolizine. Synthesis of enantiomerically pure (S)-acetoxyglutarimide, stereoselective reductive intramolecular cyclization, hydroxyl group-assisted in situ N-Boc-deprotection, selective deoxygenation of the xanthate ester, and lactam hydrolysis followed by an appropriate exchange of nitrogen regioselectivity in intramolecular cyclization were the decisive steps.

A unified synthesis of all stereoisomers of 2-(aminomethyl)cyclobutane-1-carboxylic acid

All four stereoisomers of 2-(aminomethyl)cyclobutane-1-carboxylic acid have been prepared starting from a single chirally derivatized unsaturated γ-lactam. A photochemical [2+2] cycloaddition reaction of this compound with ethylene was employed to create the four-membered ring exclusively with a cis ring configuration. The two diastereoisomers were separated conveniently by chromatography then the chiral appendage replaced by the more convenient Boc protecting group. Base-mediated lactam hydrolysis provided each of the cis target structures as single enantiomers. An efficient, tandem epimerization/hydrolysis protocol was applied to a carboxamide derivative of each cis compound, to furnish the corresponding trans compounds as single isomers, for the first time.

The Stabilizing Effect of Moisture on the Solid-State Degradation of Gabapentin

Gabapentin is known to undergo intramolecular cyclization to form a lactam (gaba-L) with concomitant loss of water. Gabapentin was milled in a planetary mill for 15-60 min. Unmilled and milled gabapentin were stored at 50°C with relative humidity ranged between 5% and 90%. The unmilled and milled samples were assayed for gabapentin and gaba-L by reversed phase-high-performance liquid chromatography and also subjected to powder X-ray diffraction, solid-state nuclear magnetic resonance and surface area analyses. The rates of lactamization in the milled gabapentin samples correlated to increased surface area, milling duration, and in-process lactam levels. This effect of milling could not be explained solely by the increase in surface area with increased milling time but was more likely due to increased regions of crystal disorder caused by the mechanical and thermal milling stresses. The lactamization rate of milled gabapentin samples was greatest in the presence of the lowest humidity conditions and dramatically decreased with increasing humidity. In particular, milled gabapentin appeared to be much more stable at humidity levels greater than 31% RH. This finding could not be attributed to the possibility of lactam hydrolysis at high humidity but rather to a competitive annealing process wherein milling-induced crystal defects were lost upon exposure to atmospheric moisture thereby stabilizing the milling-damaged drug substance.

Activation of the hydrolytic polymerization of ε-caprolactam by ester functions: Straightforward route to aliphatic polyesteramides

The hydrolytic polymerization of ε-caprolactam (CLa) was carried out in bulk (in absence of solvent) at 250 °C in the presence of carboxylic esters and aqueous H 3PO 2. It turned out that by conducting the ring opening polymerization (ROP) of CLa in the presence of PEO–C(O)–O–C 5H 11, a selected model ester (PEO = poly(ethylene oxide)), a remarkable activating effect of the ester function on the hydrolytic polymerization of the lactam was observed yielding PEO– b–PCLa diblock copolymers. The comparison of the CLa monomer conversions obtained with or without the model ester activated by H 3PO 2, as determined by 1H NMR spectroscopy, has enabled to propose a multi-step mechanism in which three major reactions occurred: (i) ester and lactam hydrolysis, (ii) aminolysis of the carboxylic ester by the resulting primary amine of the hydrolyzed/opened lactam ring and (iii) condensation reactions between carboxylic acids and both amine/hydroxyl functions. The overall result of this multi-step mechanism can be assimilated as an “insertion” of the opened lactam into the ester function. By conducting the hydrolytic polymerization of CLa in the presence of an aliphatic polyester chain, such as poly(ε-caprolactone) (PCLo), polyesteramides were recovered with high yields and random distributions of the CLa and CLo repetitive units as determined by 13C NMR.

PH gradients in immobilized amidases and their influence on rates and yields of ?-lactam hydrolysis

The pH gradients developing within immobilized biocatalysts during hydrolysis of penicillin G and glutaryl-7-aminocephalosporanic acid have been estimated both theoretically and experimentally. For the latter a fluorimetric method for the direct measurement of the average pH value within the carrier during reaction has been developed using the pH-dependent fluorescence intensity of an enzyme-bound fluorophore determined with a fiber bundle. The theoretical calculations were based on a model for the hydrolysis with immobilized enzymes using a kinetic expression with five pH-dependent, measurable kinetic and equilibrium constants. The transport reaction differential equation which considers the laminar boundary layer has been solved numerically for the key component. The calculated values agreed well with the experimental data. Under the typical reaction conditions of penicillin G hydrolysis the average pH value in the carrier was 1 and 2.5 pH units below the bulk pH (=8) with and without buffer, respectively. The corresponding changes for the hydrolysis of glutaryl-7-aminocephalosporanic acid at bulk pH 8 in the presence of buffer was 0.5. This demonstrates the existence of considerable pH gradients in carriers during hydrolytic reactions, even in buffered systems with negligible mass transfer resistance. The low pH value causes suboptimal reaction rates, reduced equilibrium conversion, and reduced enzyme stability. These pH gradients can be minimised by using buffers with pK values approximately equal to the bulk pH used for the hydrolysis. The prediction quality of the model has been tested applying it to fixed bed reactor design. The reduction in rate and yield due to concentration and pH gradients can be overcome with simple measures such as high initial pH value and pH adjustments in segmented or recycling fixed bed reactors. Thus, enzymatic conversions with high yield and high operational effectiveness are achieved.

Chiral bicyclic Lactams as homoenolate equivalents: An asymmetric route to 5-substituted cyclohexenones

Bicyclic Lactam 2 was transformed into α-cyanoenamine 8 in a four-step, high-yielding process. γ-Metalation of 8 was achieved using LiTMP in THF/HMPA, and the resultant homoenolate equivalent underwent stereoselective alkylation and subsequent hydrolysis to provide lactams of type 10 as a single diastereomer. Partial reduction and hydrolysis of lactams 10 furnished enantiopure 5-substituted cyclohexenones 13 in good overall yield.

Enantioselective Synthesis of 2-Substituted 5-, 6- and 7-Membered Lactams via α-Alkylation of Their Chiral N-Dialkylamino Derivatives

2-Alkyl-substituted lactams 4 were synthesized in good overall yields and high enantiomeric purities (ee =71-99%) by α-alkylation of chiral N-(dialkylamino)lactams 2 and subsequent reductive N-N bond cleavage of the resulting lactams 3 with lithium in liquid ammonia. The lactams 2, in turn, were prepared in good yields by cyclization of ω-chloroalkanohydrazides 1 with sodium hydride. Acidic hydrolysis of lactams 4 leads to γ-aminobutanoic acid (GABA) derivatives 5 (ee ≥ 99%). The absolute configuration was determined by polarimetry and an X-ray structure analysis of (S,S)-3e’.

The kinetics of non-stoichiometric bursts of β-lactam hydrolysis catalysed by class C β-lactamases

Class C beta-lactamases from Pseudomonas aeruginosa and several species of the Enterobacteriaceae have been observed to undergo a rapid burst in hydrolysis of beta-lactam antibiotics before relaxation to a steady-state rate of hydrolysis. The amplitude of the burst corresponds to the hydrolysis of between 1 and 10,000 mol of the substrate per mol of enzyme. The decay of the rate of hydrolysis in the burst phase comprises two exponential reactions, which indicates that there are three different reactive states of the enzymes. Examination of the kinetics of acylation by slowly reacting beta-lactams suggests that there are three forms of the free enzyme in slow equilibrium. Thus it would appear that the burst kinetics exhibited by class C enzymes can be attributed to redistribution of the enzyme between different conformations induced by the reaction with substrate.

A dramatic change in the rate-limiting step of beta-lactam hydrolysis results from the substitution of the active-site serine residue by a cysteine in the class-C beta-lactamase of Enterobacter cloacae 908R.

A cysteine residue has been substituted for the active-site serine of the class-C beta-lactamase produced by Enterobacter cloacae 908R by site-directed mutagenesis. The modified protein exhibited drastically reduced kcat./Km values on all tested substrates. However, this decrease was due to increased Km values with some substrates and to decreased kcat. values with others. These apparently contradictory results could be explained by a selective influence of the mutation on the first-order rate constant characteristic of the acylation step, a hypothesis which was confirmed by the absence of detectable acylenzyme accumulation with all the tested substrates, with the sole exception of cefoxitin.

A novel synthetic route to 6‐azabicyclo[3.2.1]oct‐3‐enes via silicon directed N‐Acyliminium chemistry: Synthesis of (±)‐gabaculine

AbstractAlkylation of the lithium enolate, derived from 5‐ethoxy‐2‐pyrrolidone 1 (Scheme 1), using alkyl halides containing either the vinylsilane or 2‐propynylsilane moiety, provides substrates 3 for cyclization studies passing through N‐acyliminium intermediates 4 toward the 6‐azabicyclo[3.2.1]‐octanes 5 and 7. These ring‐closure experiments are unsuccessful in the case of vinylsilanes, and an explanation for this failure is presented. Ring closures of 2‐propynylsilanes lead, in high yield, to bicyclic allenes 5, which, in a few steps (ozonolysis, Shapiro reaction), are transformed into 7. This methodology constitutes the basis of a total synthesis of (±)‐gabaculine (10). The synthesis is completed from 52 by lactam hydrolysis and sulfoxide elimination.

A mild two-step method for the hydrolysis of lactams and secondary amides

A mild two-step method for the hydrolysis of lactams and secondary amides

Kinetics and mechanism of hydrolysis of lactams in aqueous H2S04 solutions

1. A study has been made of the kinetic relationships in the hydrolysis ofγ-butyrolactam,δ-valerolactam,e-caprolactam, and enantholactam in 0.5–65% H2SO4 at 59.5–94°C, and an analysis has been made of the literature data for the hydrolysis ofβ-,γ-, δ-, ande-lactams and N-ethylacetamide in H2SO4, solutions. 2. The influence of the medium on the rate constants ofγ-, δ-, e-, and enantholactams and N-ethylacetamide can be explained quantitatively on the assumption that the hydrolysis proceeds along two pathways. The reaction rates are determined as follows. Along the first pathway, the rate is determined by the product of concentratrations of the un-ionized form of the reagent and H502+ ions. Along the second pathway, the rate is determined by the concentration of the complex BH30+, which consists of the form of the lactam that is protonated at the carbonyl group and a water molecule coordinated on the carbon atom of the carbonyl group. 3. Values have been calculated for the rate constants determining the contributions of the individual pathways to the experimentally determined rate constant. 4. In the hydrolysis ofβ-propiolactam, the limiting stage is the decomposition of the complex consisting of the un-ionized form of the lactam (B) and the ion pair H30+·H2SO4−. The concentration of such a complex is determined by the product of the concentration of (B) and the thermodynamic activities of the acid and water. Under conditions of hydrolysis, the relative concentration of such a complex is small.

The kinetics and mechanism of acid catalysed hydrolysis of lactams

The acid-catalysed hydrolysis of lactams of ring size 4–7, and of N-ethylacetamide, has been studied at various temperatures in 0–70% aqueous sulfuric acids. The kinetics have been analysed by means of the r-hydration parameter treatment and the transition-state activity coefficient (TSAC) approach. Except for the β-lactam, all substrates show a strong positive rate dependence on water activity, indicating a tetrahedral intermediate type of mechanism [Formula: see text], similar to that found for simple (acyclic) amide hydrolyses; only the β-lactam shows a negative rate – water activity dependence, which is interpreted in terms of a unimolecular [Formula: see text] mechanism, involving rate-determining acylium ion formation. Similarly, all substrates except the β-lactam show a pronounced destabilization of the transition-state as acidity is increased, typical of oxonium ion species. The behaviour of the β-lactam resembles that of methyl mesitoate hydrolysis. Basicity constants for lactams of ring size 4–9 show a definite dependence on ring size, with the δ-lactam being unusually basic. Both rate and pK dependence on ring size are discussed in terms of ring strain and steric effects. The activation parameters, particularly ΔS≠, are fully consistent with the conclusions drawn concerning transition-state hydration.

Mechanism of the hydrolysis of lactams in aqueous solutions of potassium hydroxide

The subject of this study was the kinetics of the hydrolysis of γ-butyrolactam, δ-valerolactam, ε-caprolactam and enantholactam in strong aqueous solutions of potassium hydroxide. It has been shown that the average thermodynamic activity of aqueous solutions of potassium hydroxide is a function of alkalinity. Ionization of lactams yields singly and doubly ionized forms of reactive lactams. The processes of interaction of these ions with the water molecule are the rate-limiting steps.

Amines and carboxylic acids as initiators of polymerization in caprolactam

AbstractEvidence is presented for the initiation of polymerization in caprolactam by two primary amines and two carboxylic acids, with rigorous precautions to exclude water from the systems. The kinetics of the reactions have been studied dilatometrically and all are shown to share two major features: (1) the plot of conversion vs. time is characteristically sigmoid in shape, and (2) the maximum slope of this plot tends to be proprtional to the square root of the initiator concentration. The rate data show that acids induce rather more rapid polymerization than bases. When combined with data in the literature on the polymerization induced by aminocaproic acid or water, they indicate that reaction of lactam at the amino and carboxyl groups accounts for an appreciable but minor proportion of the observed reaction rate; the remainder is presumably attributable to the mechanism of alternate hydrolysis of lactam and condensation of terminal amino and carboxyl groups. The explanation of the precise mechanism by which amines and acids induce polymerization offers some difficulty. A molecular ring‐addition mechanism fails to account for the square root concentration dependence mentioned above. The hypothesis of ring‐insertion at chain amide linkages is capable of yielding a suitable rate dependence but is shown to be invalidated by other evidence. A tentative suggestion of an ionic ring‐addition process is made which is consistent with the major kinetic characteristics of these reactions.