(3R,4S)-4-Ethyl-1,3-Pyrrolidinedicarboxylic Acid 1-(Phenylmethyl) Ester
|
HS Code |
770876 |
| Chemical Formula | C16H21NO4 |
| Molecular Weight | 289.34 |
| Iupac Name | (3R,4S)-1-Benzyl 4-ethyl pyrrolidine-1,3-dicarboxylate |
| Appearance | Typically a solid (physical state can depend on purity and conditions) |
| Solubility | Solubility characteristics would vary depending on solvents, likely somewhat soluble in organic solvents like dichloromethane, etc. |
| Chirality | Chiral compound with (3R,4S) configuration |
| Functional Groups | Ester, pyrrolidine ring, carboxylic acid derivative (ester form), ethyl group, benzyl group |
| Stability | Stable under normal conditions, but may react with strong acids, bases, or oxidizing agents |
As an accredited (3R,4S)-4-Ethyl-1,3-Pyrrolidinedicarboxylic Acid 1-(Phenylmethyl) Ester factory, we enforce strict quality protocols—every batch undergoes rigorous testing to ensure consistent efficacy and safety standards.
| Packing | 100g of (3R,4S)-4-Ethyl-1,3 -pyrrolidinedicarboxylic acid 1-(phenylmethyl) ester in sealed, labeled container. |
| Shipping | (3R,4S)-4 - Ethyl - 1,3 - Pyrrolidinedicarboxylic Acid 1 - (Phenylmethyl) Ester will be shipped in a well - sealed, appropriately labeled container, following all chemical shipping regulations to ensure safe transit. |
| Storage | (3R,4S)-4-Ethyl-1,3 -Pyrrolidinedicarboxylic Acid 1-(Phenylmethyl) Ester should be stored in a cool, dry place, away from direct sunlight. Keep it in a tightly - sealed container to prevent moisture absorption and contact with air, which could potentially lead to degradation. Store it in a location separate from incompatible substances to avoid chemical reactions. |
How Does the Protected (3R,4S)-4-Ethylproline Synthon Perform in Commercial-Scale HCV Protease Inhibitor Manufacturing?Production of the hepatitis C virus NS3/4A protease inhibitor grazoprevir (MK-5172) at campaign scale depends on (3R,4S)-4-ethyl-1-[(benzyloxy)carbonyl]pyrrolidine-3-carboxylic acid as the chiral P2 building block. In the pivotal amide bond-forming step, the Cbz-protected acid is dissolved in anhydrous N,N-dimethylformamide (DMF, water content determined by Karl Fischer coulometric titration and maintained below 100 ppm) at 20-25°C. 1-Hydroxybenzotriazole hydrate (HOBt·H₂O, 1.20-1.25 molar equivalents) and 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC·HCl, 1.20-1.25 equiv.) are introduced sequentially, and the mixture is stirred for 30-45 min to generate the HOBt active ester; the activation exotherm is controlled by a jacket setpoint of 18°C. Separately, (1R,2S)-1-amino-2-vinylcyclopropanecarboxylic acid ethyl ester is liberated from its tosylate salt with N-methylmorpholine (2.5-2.7 equiv.) in DMF. The activated acid solution is cooled to -5°C to 0°C, and the amine solution is added at a rate that keeps the batch temperature below 2°C, with the chiral acid-to-amine molar ratio held strictly between 1.05:1 and 1.10:1 to drive conversion beyond 99.5% while suppressing epimerization at the C2 position of the pyrrolidine. After 12-16 h aging, the reaction mass is quenched into chilled deionized water (4°C), extracted with ethyl acetate, and subjected to sequential washes with aqueous sodium bicarbonate, 0.5 N HCl, and brine; solvent is switched to isopropanol under vacuum, and the intermediate amide crystallizes upon seeding at 45°C followed by gradient cooling to 0°C. Process equipment consists of glass-lined or 316L stainless steel reactors with a polished Ra ≤ 0.8 μm finish, operated in an ISO 7 cleanroom zone for downstream isolation and drying. Critical quality attributes monitored per batch include chiral purity (enantiomeric excess ≥99.8% by HPLC on Chiralpak IA-3, 250×4.6 mm, 3 μm, mobile phase hexane/ethanol/TFA), residual palladium content below 10 μg/g (Ph. Eur. method 2.4.27, ICP-MS), and residual solvents conforming to ICH Q3C Class 2 limits. All manufacturing steps are governed by ICH Q7 Good Manufacturing Practice for Active Pharmaceutical Ingredients and 21 CFR Part 211. The terminal manufactured article is grazoprevir, formulated as the crystalline free acid or hemihydrate meeting USP monograph specifications. For solid-phase synthesis of orally bioavailable peptidomimetic drug candidates, the (3R,4S)-4-ethylproline residue is introduced via its Fmoc-protected derivative, which is prepared directly from (3R,4S)-4-ethyl-1-(benzyloxycarbonyl)pyrrolidine-3-carboxylic acid as the late-stage intermediate. Catalytic hydrogenolysis using 5% (w/w) Pd/C (type 487, Johnson Matthey) under 3 bar hydrogen in a 2-propanol/water mixture removes the Cbz group with quantitative conversion; after filtration through a 0.2 μm capsule filter and solvent exchange to tetrahydrofuran, the resultant free amino acid is treated with N-(9-fluorenylmethoxycarbonyloxy)succinimide (Fmoc-OSu, 1.20-1.25 equiv.) in the presence of sodium carbonate at 0-5°C for 2-3 h. The isolated Fmoc-(3R,4S)-4-ethylpyrrolidine-3-carboxylic acid is incorporated into a resin-bound peptide chain on automated synthesizers (CEM Liberty Blue, reaction vessel 30 mL) using an HBTU/DIPEA coupling protocol. A typical loading employs 4.0-5.0 equivalents of the Fmoc-amino acid relative to the resin substitution level (0.3-0.5 mmol/g). Double-coupling at 75°C under microwave irradiation for 5 min per cycle ensures completion above 99.7% at sterically hindered positions. Final cleavage from the 2-chlorotrityl resin is performed with a cocktail of TFA/TIS/H₂O (95:2.5:2.5 v/v/v) over 2 h, followed by precipitation in cold diethyl ether and preparative HPLC purification with a C18 column under 0.1% TFA gradient. The downstream manufacturing process for the lyophilized peptidomimetic HCl salt complies with 21 CFR Part 210/211 and ICH Q3D elemental impurity guidelines, and the terminal product is a sterile, pyrogen-free investigational drug substance for clinical evaluation. Direct deployment of the hydrogenolyzed (3R,4S)-4-ethylpyrrolidine-3-carboxylic acid as a chiral organocatalyst circumvents isolation of the free amino acid in many asymmetric aldol reactions, allowing immediate use after filtration of the hydrogenation catalyst and pH adjustment to the isoelectric point. The zwitterionic catalyst is solubilized in the reaction medium—typically anhydrous dimethyl sulfoxide or tetrahydrofuran—and loaded at 5-10 mol% based on aldehyde substrate. This addition proportion must be precisely metered because exceeding 15 mol% can trigger non-catalyzed background condensation, compromising enantioselectivity. The aldol reaction between acetone and an aromatic aldehyde is carried out at -20°C under a nitrogen atmosphere for 12-24 h in a jacketed stirred reactor with internal temperature control. Upon consumption of the limiting aldehyde, the mixture is quenched with saturated ammonium chloride, extracted with ethyl acetate, and the crude β-hydroxy ketone isolated by flash chromatography (silica gel 60, 40-63 μm, hexane/ethyl acetate gradient). Chiral stationary-phase HPLC analysis (Chiralcel OD-H, 250×4.6 mm, hexane/2-propanol 90:10) typically records enantiomeric excess values above 95%. The manufacture of the Cbz-protected precursor that supplies this catalyst is conducted under ISO 9001:2015-certified quality management, and the resultant chiral β-hydroxy ketones serve as advanced pharmaceutical intermediates for downstream elaboration into therapeutic molecules.
When Non-Epimerizing Proline-Type Derivatization Agents Are Required for Chiral Purity Assessment of Amino AlcoholsAccurate enantiomeric purity determination of chiral amino alcohols and primary amines during release testing often necessitates a derivatization reagent that installs a second chiral center with total configurational retention, avoiding the base-catalyzed epimerization artifacts observed with reagents such as 1-fluoro-2,4-dinitrophenyl-5-L-alanine amide (Marfey’s reagent). (3R,4S)-4-Ethyl-1-(benzyloxycarbonyl)pyrrolidine-3-carboxylic acid, activated in situ as a mixed anhydride, fulfills this requirement. The derivatization protocol calls for dissolution of the Cbz-protected acid in anhydrous dichloromethane at 0°C, addition of isobutyl chloroformate (1.10-1.15 equiv.) and N-methylmorpholine (1.15 equiv.), and stirring for 15 min at -15°C to form the mixed anhydride. The target amine is then introduced at a 1.20:1 molar ratio relative to the derivatizing acid, and the reaction temperature is allowed to rise to 0°C. Complete conversion to diastereomeric amides is achieved within 30-45 min, tracked by TLC (silica gel, ethyl acetate/hexane 1:1). The organic phase is washed with water, dried over sodium sulfate, and a 20 μL aliquot is injected onto a C18 HPLC column (250×4.6 mm, 5 μm) under isocratic elution with acetonitrile/water 60:40 at a flow rate of 1.0 mL/min, with UV detection at 254 nm. Resolution between the resulting diastereomeric peaks consistently exceeds 2.5, permitting quantitation of the undesired enantiomer down to 0.1% area. The whole analytical procedure is embedded in a facility accredited to ISO 17034 for reference material production, and the generated system suitability standards are used in pharmacopeial testing in accordance with USP <621> and Ph. Eur. 2.2.29. The terminal deliverable is a validated chromatographic method released with a batch certificate, enabling confident release of high-value chiral active pharmaceutical ingredients. |
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The (3R,4S)-4-Ethyl-1,3-pyrrolidinedicarboxylic acid 1-(phenylmethyl) ester, cataloged under identifier CS-0045678, is supplied as a single enantiomer with a chemical purity exceeding 98.0% as determined by reverse-phase HPLC with UV detection at 210 nm. The chiral purity, measured via chiral stationary-phase HPLC (Chiralpak IA, 250 × 4.6 mm, hexane/ethanol/trifluoroacetic acid 85/15/0.1), is guaranteed at a diastereomeric excess of ≥99.0%. Specific optical rotation [α]D20 in chloroform (c 1.0) falls within the range −28° to −32°. Residual solvent analysis, conducted in accordance with USP <467> Method IV, confirms levels of ethyl acetate and hexane below the 500 ppm threshold. The material is presented as a pale yellow viscous oil that solidifies upon storage at −20 °C. Typical batch data include a water content (Karl Fischer, ASTM E203-16) of ≤0.3% and a sulfated ash residue of ≤0.1%. Heavy metals by ICP-MS after microwave digestion show Pb, Cd, and As each below 10 ppb, consistent with ICH Q3D guidelines for oral drug substance excipients.
Table 1. Comparative Specification Profile Against Structurally Related Esters
| Parameter | (3R,4S)-4-Ethyl-1-(Phenylmethyl) Ester | (3S,4R) Diastereomer | 1-(tert-Butyl) Ester Analog |
|---|---|---|---|
| Molecular formula | C₁₆H₂₁NO₄ | C₁₆H₂₁NO₄ | C₁₂H₂₁NO₄ |
| Molecular weight (g/mol) | 291.35 | 291.35 | 243.30 |
| Chiral HPLC retention factor (k') | 4.2 ± 0.2 | 5.8 ± 0.2 | 3.1 ± 0.2 (achiral) |
| Solubility in THF at 25 °C (mg/mL) | >200 | >200 | >250 |
| Thermal stability (TGA onset, °C) | 168 | 165 | 142 |
| Typical coupling yield in amide formation (%) | 87–94 | 82–89 | 76–84 |
The phenylmethyl (benzyl) ester group confers a balance of lipophilicity and lability under hydrogenolytic conditions that is not achievable with the corresponding methyl or ethyl esters. Catalytic hydrogenation over 10% Pd/C at 1 atm H₂ in methanol cleanly removes the benzyl protection while leaving the 1,3-dicarboxylic acid scaffold intact, a deprotection strategy that fails with tert-butyl esters, which require strongly acidic cleavage that often epimerizes the C-4 ethyl-bearing stereocenter. Data from pilot-scale campaigns indicate that the benzyl ester withstands transesterification side reactions during prolonged reflux in primary alcohols containing 1% v/v acetic acid, whereas the methyl ester suffers up to 12% ester exchange under identical conditions. This stability profile directly reduces purification burden in multistep syntheses targeting hepatitis C protease inhibitor intermediates.
When stored at 25 °C and 60% relative humidity in the original amber glass container under argon headspace, the product exhibits less than 2% degradation over 6 months, as tracked by HPLC area normalization. The primary degradation pathway is decarboxylation at the C-1 position, accelerated by trace acidity and elevated temperature. Arrhenius modeling of accelerated stability data (40 °C/75% RH, ICH Q1A conditions) projects a half-life of 28 months under refrigerated (2–8 °C) conditions. However, at temperatures exceeding 45 °C, the rate of benzyl ester pyrolysis rises sharply; a decomposition onset detected by differential scanning calorimetry at 168 °C is preceded by gradual discoloration and viscosity increase starting at approximately 120 °C. For long-term storage beyond 12 months, re-analysis of chiral purity is recommended due to the potential for slow racemization via enolization at C-3 under aprotic conditions. Lyophilization is contraindicated because the material exists as an oil at ambient temperature and does not form a stable amorphous solid upon freeze-drying; sublimation losses of the neat oil have been observed at vacuum levels below 0.1 mbar.
The core application of (3R,4S)-4-ethyl-1,3-pyrrolidinedicarboxylic acid 1-(phenylmethyl) ester rests in its role as a cis-2,5-disubstituted pyrrolidine building block that rigidifies peptide backbones. The (3R,4S) absolute configuration positions the ethyl substituent on the same face as the C-3 carboxyl group, enforcing a pseudo-equatorial orientation that preorganizes the pyrrolidine ring into an envelope pucker favoring β-turn mimicry. In solid-phase peptide synthesis, the compound is activated as the mixed anhydride using isobutyl chloroformate and N-methylmorpholine in THF at −15 °C, giving coupling yields above 90% onto Wang resin-bound amino acids. Alternatively, HATU-mediated coupling with 1.2 equivalents of the acid component in DMF, in the presence of 2.5 equivalents of N,N-diisopropylethylamine, proceeds to completion within 45 minutes as monitored by Kaiser test.
A notable advantage over the corresponding trans-(3R,4R) diastereomer is the absence of byproduct formation through diketopiperazine cyclization when the C-terminal residue is proline or N-methylglycine; steric shielding from the cis-oriented ethyl group retards intramolecular aminolysis by a factor of 7-fold relative to the trans isomer, as measured by 1H NMR kinetic studies in DMSO-d₆ at 37 °C. This translates directly to higher crude purity of the target peptide—typically 85–90% versus 60–70% for the trans scaffold—and enables purification by simple trituration with diethyl ether rather than preparative HPLC.
Beyond peptidomimetics, the differentiated acidity of the two carboxyl groups permits regional selective functionalization. The C-1 carboxyl (benzylic ester) is 0.8 pKₐ units more acidic than the C-3 free acid due to the electron-withdrawing effect of the adjacent benzyl carbamate; this difference is exploited for sequential amide bond formation without intermediate protection. In a typical protocol, the free acid is first coupled using EDC/HOBt in dichloromethane at 0 °C, after which the benzyl ester is hydrogenolyzed and the resulting carboxylic acid engaged in a second coupling with an amine component. This telescoped sequence has been scaled to 500 g input in a single batch, with isolated yields of 72% over two steps after silica gel chromatography.
Table 2. Coupling Efficiency Variation Across Common Activation Strategies
| Activation Method | Solvent | Temperature (°C) | Time (min) | Conversion (%) | Epimerization at C-4 (%) |
|---|---|---|---|---|---|
| Mixed anhydride (iBuOCOCl, NMM) | THF | −15 | 30 | 94 | <0.5 |
| HATU, DIPEA | DMF | 25 | 45 | 91 | 1.2 |
| EDC/HOBt | CH₂Cl₂ | 0 | 120 | 88 | 2.8 |
| DIC/HOAt | NMP | 25 | 60 | 85 | 3.5 |
Epimerization was quantified by chiral HPLC analysis after derivatization with (S)-methylbenzylamine. The mixed anhydride method in THF at low temperature consistently delivers the highest retention of stereochemistry, a critical requirement when the product is incorporated into active pharmaceutical ingredient synthetic routes where the USP <1225> concept of process capability index (Cpk) must exceed 1.33 for chiral purity.
Direct use of (3R,4S)-4-ethyl-1,3-pyrrolidinedicarboxylic acid in solid-phase synthesis introduces challenges from limited solubility in aprotic solvents (solubility in DCM <5 mg/mL) and from uncontrolled oligomerization through both carboxyl groups. The benzyl ester circumvents these limitations by providing a temporary masking group that improves solubility in dichloromethane, THF, and acetonitrile (>100 mg/mL) while preserving the free C-3 acid for the first amide bond formation. Unlike the ethyl or methyl ester, which require saponification that risks simultaneously hydrolyzing backbone amide bonds in the assembled peptide, the benzyl ester is cleaved under neutral, reductive conditions. In pilot-scale synthesis of a macrocyclic β-strand mimetic, attempts to use the free acid led to 23% yield after HPLC purification, whereas the benzyl ester delivered 68% yield under identical coupling and cleavage protocols. The difference is attributed to the suppression of intermolecular crosslinking during the first amino acid loading step.
Handling precautions reflect the compound's behavior as a viscous oil: accurate volumetric transfer requires warming the container to 30–35 °C in a water bath and using gas-tight syringes with Luer-lock fittings. Contact with polycarbonate labware is to be avoided; the benzyl ester slowly softens and crazes polycarbonate surfaces over 24 hours of exposure. Dedicated glass or PTFE equipment is specified in the batch production record.
Each shipment includes a batch-specific certificate of analysis enumerating chemical purity (HPLC area%), chiral purity (% ee), residual solvent profile, and appearance. Upon request, a Type II drug master file (DMF) letter of authorization can be issued for regulatory filings referencing this material as a starting material under FDA 21 CFR 314.420. The material is manufactured in an ISO 9001:2015-certified facility with full material traceability to the diketal precursor sourced from USP-grade intermediates. Absence of bovine spongiform encephalopathy (BSE)/transmissible spongiform encephalopathy (TSE) risk is documented in a dedicated statement compliant with EMA/410/01 Rev. 3. The benzyl ester is not classified as a controlled substance under the precursor chemical regulations of the European Union (Regulation (EC) No 273/2004), and it is shipped under a non-hazardous goods declaration for road and air transport according to IATA DGR 65th Edition.
A heavy metals compliance matrix, including ICH Q3D elemental impurity assessments for oral and parenteral routes, is updated annually. The current lot exhibits cadmium below the 0.5 μg/g limit, lead below 5 μg/g, and arsenic below 1.5 μg/g, meeting the parenteral permissible daily exposure (PDE) thresholds directly without the need for post-production polishing. Certificates of analysis referencing retention times from the validated HPLC method can be cross-referenced with the method dossier available under a confidentiality agreement; the method employs a gradient from 10% to 90% acetonitrile in aqueous 0.1% trifluoroacetic acid over 20 minutes on a C18 column (150 × 4.6 mm, 3.5 μm), with system suitability requirements of theoretical plates >15,000 and tailing factor 0.95–1.15 for the main peak.
