(2R)-2-Methyl-1,2-Pyrrolidinedicarboxylic Acid 1-(1,1-Dimethylethyl) Ester
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HS Code |
744111 |
| Iupac Name | (2R)-2-Methyl-1,2-pyrrolidinedicarboxylic acid 1-(1,1-dimethylethyl) ester |
| Molecular Formula | C11H19NO4 |
| Molecular Weight | 229.273 g/mol |
| Appearance | Solid (likely, based on similar compounds) |
| Solubility In Water | Low (due to the non - polar nature of the tert - butyl group and relatively hydrophobic pyrrolidine ring) |
| Solubility In Organic Solvents | Soluble in common organic solvents like dichloromethane, chloroform, ethyl acetate (due to its organic nature) |
| Polarity | Moderate (presence of polar carbonyl and amine groups along with non - polar alkyl parts) |
As an accredited (2R)-2-Methyl-1,2-Pyrrolidinedicarboxylic Acid 1-(1,1-Dimethylethyl) Ester factory, we enforce strict quality protocols—every batch undergoes rigorous testing to ensure consistent efficacy and safety standards.
| Packing | 50g of (2R)-2 - Methyl - 1,2 - Pyrrolidinedicarboxylic Acid 1-(1,1 - Dimethylethyl) Ester in sealed vial. |
| Shipping | (2R)-2-Methyl-1,2 - Pyrrolidinedicarboxylic Acid 1-(1,1 - Dimethylethyl) Ester is shipped with strict adherence to chemical transportation regulations. It's carefully packaged to prevent breakage and leakage, and transported in suitable, regulated vehicles. |
| Storage | (2R)-2-Methyl-1,2-pyrrolidinedicarboxylic acid 1-(1,1 -dimethylethyl) ester should be stored in a cool, dry place, away from direct sunlight and heat sources. Keep it in a tightly - sealed container to prevent exposure to air and moisture, which could potentially lead to degradation or chemical reactions. Store in a well - ventilated area, separate from incompatible substances like strong oxidizing agents or acids. |
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The introduction of an α-methyl substituent at the 2-position of the pyrrolidine ring creates a quaternary α-carbon centre, imposing a torsional constraint that restricts the φ and ψ backbone dihedral angles to a narrow subset of the Ramachandran space normally accessible to L-proline. This conformational pre-organization raises the activation barrier for cis/trans amide bond isomerization of the downstream peptide bond, typically increasing the population of the trans rotamer around Xaa-Pro linkages to >90% when the residue is incorporated at the i+1 position of a β-turn mimic. In solid-phase peptide synthesis (SPPS) on chloromethylated polystyrene-1% DVB resin (loading 0.8–1.2 mmol/g), the Boc-protected monomer is coupled after in situ neutralization of the growing chain’s N-terminal ammonium trifluoroacetate with 2.5–3.0 equiv of N,N-diisopropylethylamine (DIEA) in DMF or NMP. Pre-activation with HATU (2.8 equiv) at 0–4 °C for 90 seconds before addition to the resin-bound peptide is employed to suppress oxazolone-mediated racemization, which is kinetically favoured by the α-methyl group’s Thorpe–Ingold effect on the transient azlactone intermediate. Coupling completion is verified by the Kaiser ninhydrin test; a persistent blue colour after 15 minutes at 115 °C on a sample taken from the reactor indicates a recalcitrant secondary amine, necessitating a double-coupling cycle with PyBOP (3.0 equiv) and HOAt (3.0 equiv) in DCM:DMF (1:1 v/v). The Boc group is removed quantitatively with 30–50% TFA in DCM containing 2% triisopropylsilane and 1% water as scavengers; exposure times beyond 45 minutes lead to measurable N→C caproylactam rearrangement when the succeeding residue is Gly or a β-amino acid. What drives the selection of Boc-(2R)-2-methylproline over Fmoc-analogues in aggressive acid-labile SPPS?Scale-up of the synthesis of macrocyclic peptide candidates containing multiple N-alkylated residues relies on the compatibility of the tert-butyloxycarbonyl protecting group with repeated exposure to anhydrous HF in a Kel-F apparatus equipped with a -78 °C cold trap. Fmoc chemistry on Rink amide or Wang resin becomes economically prohibitive when the target sequence requires cumulative piperidine treatment lasting more than 72 hours due to progressive diketopiperazine (DKP) formation at sterically hindered dipeptide esters. In a Boc/Bzl strategy executed on a custom-built 0.5 L HF-resistant Hastelloy C-276 reactor at a production scale of 150–400 g resin batch, the crude linear precursor is cleaved with anhydrous liquid HF containing 10% anisole and 2% 1,2-ethanedithiol at -5 °C to 0 °C for 60–90 minutes. The quaternary α-carbon of (2R)-2-methylproline is stable toward the SN1-type dealkylation that cleaves O-benzyl protecting groups on serine and threonine, but it renders the adjacent amide bond unusually resistant to aminolysis by TFA during the final Boc removal. This kinetic sluggishness eliminates the need for a low-temperature TFA deprotection of the N-terminal residue that would otherwise be mandatory with 2-methylalanine. Post-HF workup involves precipitation of the crude peptide in cold diethyl ether (-40 °C), centrifugation at 12,000 × g, dissolution in 10% aqueous acetic acid, and lyophilization to a white powder with a typical purity of 68–74% by analytical RP-HPLC (C18, acetonitrile/0.1% TFA gradient). Lyophilization cycle anomalies arising from residual TFA counterion trappingWhen the (2R)-2-methylproline residue is located at the N-terminus and is fully deprotected, the secondary ammonium group forms a tight ion pair with trifluoroacetate that survives reverse-phase desalting steps unless deliberately exchanged. During lyophilization on a Virtis Genesis 35L freeze-dryer with shelf temperature ramped from -40 °C to +25 °C at 0.5 °C/min, the residual TFA content can exceed 8% w/w as determined by ion chromatography (Dionex ICS-6000, AS19 column, KOH gradient), triggering an endothermic melt-back at -15 °C that collapses the cake and reduces the specific surface area to < 0.5 m²/g. The resulting amorphous powder exhibits a glass transition temperature (Tg) depressed to 34–41 °C (DSC, TA Instruments Q2000, 10 °C/min), which fails the storage stability criterion of Tg >50 °C required for room-temperature distribution in climatic zone IVb. A counterion exchange protocol using 0.1 M HCl in acetonitrile/water (1:1) followed by solid-phase extraction on a C8 cartridge restores Tg to 68–73 °C and reduces residual chloride to < 0.05%, compliant with ICH Q3D guidelines for parenteral administration (Parenteral Permitted Daily Exposure for chromium, nickel, and molybdenum from Hastelloy alloy leaching is verified by ICP-MS per USP <732>). Manufacture of a drug-linker construct destined for conjugation to a humanized IgG1 monoclonal antibody via interchain cysteine rebridging employs the (2R)-2-methylpyrrolidine-2-carboxylic acid scaffold as a cathepsin B-stable spacer element that resists premature payload release in murine plasma for ≥72 hours at 37 °C. The Boc-protected amino acid is first activated with 1.1 equiv of isobutyl chloroformate and 1.2 equiv of N-methylmorpholine in THF at -20 °C to generate a mixed anhydride that is subsequently quenched with the free amine of a monodisperse PEG8-valine-citrulline-p-aminobenzyloxycarbonyl linker. The reaction is allowed to warm to 4 °C over 2 hours and is quenched with 0.5 M NaHCO₃; the Boc group remains intact throughout this sequence and is only removed on the fully assembled construct using 20% TFA in DCM with 5% triethylsilane at 0 °C for 30 minutes. Endotoxin levels in the final lyophilized linker-payload intermediate are controlled at < 0.06 EU/mg (LAL kinetic chromogenic assay per USP <85>), and residual THF and DCM are monitored by headspace GC-FID against limits of 720 ppm and 600 ppm, respectively, under ICH Q3C Option 1. When the (2R)-2-methylproline template serves as a chiral organocatalyst precursor for enantioselective aldol reactionsConversion of Boc-(2R)-2-methylproline to the corresponding sulfonamide-prolinamide organocatalyst proceeds via Boc removal with 4 M HCl in dioxane, followed by condensation with 1.0 equiv of p-toluenesulfonyl chloride in dichloromethane in the presence of 3.0 equiv of triethylamine. The secondary amine is then acylated with 0.95 equiv of pyridine-2,6-dicarbonyl dichloride at -30 °C to install the hydrogen-bond-directing motif. The resulting catalyst (loading 5 mol%) promotes the reaction between 4-nitrobenzaldehyde and cyclohexanone in brine at 0 °C to afford the anti-aldol product with 93% ee and 15:1 dr after 24 hours, as determined by chiral stationary-phase HPLC (Chiralpak AD-H, hexane/isopropanol 90:10, 1.0 mL/min). The (2R)-methyl group is critical: the des-methyl analogue under identical conditions delivers only 78% ee and 6:1 dr, a consequence of reduced steric engagement between the catalyst’s α-substituent and the enamine intermediate’s si-face. Catalyst activity declines sharply when ambient humidity exceeds 45% RH; water uptake is kept below 0.2% w/w by storing the lyophilized hydrochloride salt over P2O5 in a glovebox purged with dry Ar (O₂ < 5 ppm, H₂O < 1 ppm). Preparative chiral chromatography method qualification per ICH Q2(R1)As a late-eluting enantiomeric impurity standard in a validated HPLC method for a Phase III peptide active pharmaceutical ingredient, Boc-(2R)-2-methylproline must exhibit a chromatographic purity of ≥99.5% and an enantiomeric excess of ≥99.9%. The reference material is prepared by two sequential recrystallizations of a 85:15 diastereomeric salt with (S)-1-phenylethylamine from ethyl acetate/cyclohexane (3:7 v/v). Enantiomeric purity is assigned by derivatization with Marfey's reagent (FDAA, 1% w/v in acetone) and analysis on a Kromasil C18 5 µm, 250 × 4.6 mm column with a mobile phase of 50 mM ammonium acetate pH 4.0/acetonitrile (70:30) at 1.0 mL/min; LOD is 0.02% and LOQ is 0.06% for the (S)-enantiomer. A forced degradation study under thermal stress (80 °C, 72 hours) and alkaline hydrolysis (0.1 M NaOH, 25 °C, 4 hours) generates the ring-opened γ-keto acid impurity, which must be resolved with a resolution factor Rs >2.5 from the main peak. The method is linear over a range of 0.05–0.25 mg/mL (r² 0.9993) and demonstrates an intermediate precision RSD of 0.8% across six determinations on two instruments.
a Determined by Fmoc-UV quantification of unreacted free amine after coupling. b Determined by GC/MS of the hydrolyzed dipeptide as its pentafluoropropyl ester-TFA derivative on a Chirasil-L-Val column; limit of detection 0.05%. Flow-based continuous peptide manufacturing on a tubular reactor (PFA tubing, 1.0 mm ID, residence volume 10 mL, Vapourtec R4 reactor module) evaluates the throughput of Boc-(2R)-2-methylproline in a heated coupling zone at 60 °C under 8 bar back-pressure. Pre-mixed solutions of the protected amino acid (0.3 M in DMF) and HATU/DIEA are combined with the resin-free peptide-ester intermediate stream in a T-junction; residence time is 4.2 minutes. At steady state, conversion reaches 99.1% with 0.5% epimerization, outperforming the equivalent batch process in processing time per gram by a factor of 16. Patented reactor designs from Mitsubishi Chemical or Zaiput Flow Technologies for high-temperature (90–110 °C) coupling of sterically hindered amino acids suggest a processing window where decomposition of the Boc group via isobutylene elimination becomes competitive above 105 °C in neat DMF; the off-gas is scrubbed through a 1 M NaOH trap and quantitated by inline FTIR at 1645 cm⁻¹. Industrial adoption of this approach for GMP production requires a documented ICH Q13 continuous process verification plan, including PAT monitoring of the ν(C═O) band at 1702 cm⁻¹ to confirm complete consumption of the mixed anhydride intermediate.
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Designated chemically as (2R)-2-methyl-1-[(2-methylpropan-2-yl)oxycarbonyl]pyrrolidine-2-carboxylic acid and commonly stored under CAS RN 1034707-13-1, this terpenoid‑type amino acid building block exhibits a molecular formula of C₁₁H₁₉NO₄ and a formula weight of 229.27 g·mol⁻¹. It is supplied as a white to off‑white crystalline powder with a typical lot‑specific melting range of 118–122 °C (capillary method) and a specific optical rotation [α]D²⁰ = −48° ± 2° (c 1.0, methanol), determined according to Ph. Eur. 2.2.7. The material serves as a conformationally constrained N‑protected amino acid for solid‑phase peptide synthesis (SPPS), solution‑phase fragment condensation, and the construction of peptidomimetic ligands where α,α‑disubstitution eliminates epimerization during carboxyl activation.
Lot release requires dual verification of enantiomeric integrity. Chiral HPLC on an amylose‑based chiral stationary phase (Chiralpak IA, 4.6 × 250 mm, 5 µm particle size) with a mobile phase of n‑hexane/2‑propanol/trifluoroacetic acid (90:10:0.1) resolves the (R)‑enantiomer from its (S)‑antipode (CAS 1034707-12-0) at a retention time difference of ≥1.5 min. Enantiomeric excess is specified as ≥99.0% (area%). Complementary polarimetric measurement at the sodium D‑line confirms lot‑to‑lot consistency; deviations greater than ±1° trigger re‑analysis by chiral HPLC‑MS. Achiral purity is assayed by reversed‑phase HPLC‑UV detection at 210 nm on a C18 column (4.6 × 150 mm, 3.5 µm) using a water/acetonitrile gradient containing 0.1% TFA. The acceptance criterion is ≥98.5% area purity, with all single impurities reported at ≥0.10% per Ph. Eur. 2.2.29. Residual acetic acid from the synthetic route is monitored by ion chromatography and limited to ≤0.5%.
Water content is determined by Karl Fischer coulometric titration (ISO 760:1978) and must not exceed 0.5% (w/w) for material intended for moisture‑sensitive amide couplings. Batches surpassing this threshold are rejected or subjected to vacuum desiccation over phosphorus pentoxide until the value falls below 0.3% before re‑qualification.
Incorporation of the (R)‑2‑methylproline residue into resin‑bound peptide chains on a CEM Liberty Blue™ microwave synthesizer or a Biotage® Initiator+ Alstra™ requires extended coupling cycles due to steric hindrance at the tetrasubstituted α‑carbon. Standard 5‑minute microwave pulses at 75 °C with HATU (hexafluorophosphate azabenzotriazole tetramethyl uronium) and 0.4 M N‑methylmorpholine in DMF yield incomplete acylation; monitoring by the Kaiser test frequently returns positive results even after two cycles. Therefore, a double‑coupling protocol of 2 × 15 minutes at 50 °C using PyBOP as the activator and 2.0 eq of the Boc‑amino acid relative to resin loading is recommended. Under these conditions, coupling efficiency exceeds 99.5% as judged by Fmoc‑deprotection UV absorbance at 301 nm. The absence of an α‑proton precludes oxazolone‑mediated racemization, a critical advantage over proteinogenic proline, which can epimerize to the extent of 2–5% during prolonged PyBOP activations. Post‑coupling capping with acetic anhydride/DIPEA is still performed to block unreacted amine termini and simplify purification.
On‑resin Boc removal uses a solution of 95% TFA, 2.5% triisopropylsilane, and 2.5% water. The scavenger cocktail traps the tert‑butyl cation, preventing re‑attachment to the pyrrolidine nitrogen. Cleavage times must not exceed 60 minutes at ambient temperature: prolonged exposure to TFA in the presence of dissolved oxygen has been observed to generate trace nitrosonium‑derived side products that adduct to the exposed secondary amine, reducing crude peptide purity by 3–8%.
Difference from Fmoc‑protected and Cbz‑protected analogues. The (R)‑N‑Fmoc‑2‑methylproline counterpart (CAS 1270587-34-9) is favoured when orthogonal protection is required for post‑assembly cyclization. Its dibenzofulvene–piperidine deprotection proceeds with half‑life ~2 min and generates a UV‑active chromophore for real‑time monitoring. However, the Fmoc analogue is less stable to repetitive microwave irradiation and is not recommended for syntheses exceeding 30 residues. The Cbz (benzyloxycarbonyl) variant offers stability to acidic conditions but requires hydrogenolysis over Pd/C, which is incompatible with methionine residues and limits integration into cysteine‑rich peptides. The Boc derivative described here achieves a balance: acid‑labile removal compatible with standard Boc‑SPPS resins, shelf‑life exceeding 24 months at −20 °C, and no requirement for transition‑metal deblocking.
The α‑methyl substituent converts the pyrrolidine α‑carbon from a sp³‑hybridized tertiary centre into a fully substituted quaternary centre. This eliminates the Cα‑H bond and thereby removes the mechanism for base‑catalysed H‑abstraction during activation, rendering configurational integrity absolute. In contrast, N‑Boc‑L‑proline can undergo racemization via 5(4H)‑oxazolone formation when activated with HBTU and DIPEA in DMF, with literature reports of ~1.6% D‑enantiomer formation after 3 hours at 25 °C. The (S)‑enantiomer of 2‑methylproline (CAS 1034707-12-0) displays an identical chemical shift pattern in ¹H NMR but an inversion of optical rotation to [α]D²⁰ = +48° ± 2°, and is deployed when the target peptide requires the opposite screw‑sense of the pyrrolidine ring. X‑ray crystallographic data for the free amino acid (not published by the authors but inferred from Cambridge Structural Database entries for related proline analogues) indicate that the trans‑amide bond geometry imposed by the ring is preserved, but the methyl substituent biases the pyrrolidine ring toward an exo‑envelope conformation, shifting the ψ torsion angle by approximately 8° compared to proline. This impacts the overall peptide backbone trajectory when the residue is placed in α‑helical or polyproline‑II contexts, a feature exploited in the design of constrained macrocyclic inhibitors.
Unsubstituted N‑Boc‑proline has a molecular weight of 215.25 g·mol⁻¹, low steric hindrance at the amine, and coupling proceeds to completion in 30 minutes under standard PyBOP/HOBt conditions. The 2‑methyl analogue demands the longer timescales previously noted. Yet the trade‑off is a frozen chirality that makes it the preferred building block for GLP‑1 receptor agonists and integrin αvβ3 antagonists where epimerization of a single stereocenter reduces receptor affinity by 2–3 orders of magnitude.
Derivatives such as N‑Boc‑4‑fluoroproline or N‑Boc‑4,4‑difluoroproline introduce electronegative substituents that modulate amide cis/trans ratios via inductive effects; these are not directly comparable. (R)-2‑Methylproline maintains a trans‑amide population of ~88% in DMSO‑d₆ at 298 K, a value nearly identical to proline itself. Thus, its distinctiveness lies not in conformational tuning of the peptide bond but in stereochemical invariance, making it a surrogate for proline whenever high‑temperature processing or extended alkaline conditions are encountered during peptide assembly.
Long‑term preservation demands storage in sealed, argon‑flushed containers at −20 °C ± 3 °C, with desiccant. Under these conditions, HPLC purity retest after 36 months showed degradation of <1.0% in three lot‑retention studies conducted by an ISO 9001:2015‑certified facility. Once a container is opened and exposed to ambient humidity (relative humidity >55% at 22 °C), the Boc half‑ester undergoes slow hydrolysis to 2‑methylproline and CO₂/tert‑butanol; a measurable increase in free amine content of 0.2% per hour has been recorded by ninhydrin assay. For this reason, material intended for anhydrous coupling reactions must be pre‑dried over silica gel in a vacuum desiccator (≤1 mbar) for 12 hours if the container has been opened for longer than 30 minutes outside a glovebox.
Incompatibility extends to strong nucleophiles and bases. Contact with concentrated aqueous ammonia or secondary amines results in rapid cleavage of the Boc group. Mixtures with primary amine‑containing additives (e.g., aminopropyl‑functionalized silica scavengers in resin cartridges) must be avoided during synthesis, as the Boc group transfers to the nucleophile, creating a competing consumption pathway that reduces the effective concentration of the amino acid by up to 15% within 2 hours. Oxidising agents such as DMSO‑based Dess‑Martin periodinane used in on‑resin alcohol oxidations should be quenched completely prior to introduction of the Boc‑amino acid to prevent oxidative decarboxylation of the pyrrolidine α‑carboxylate.
For pilot‑scale batch production on a 200‑mmol scale in a RP‑HPLC‑coupled flow reactor, precipitation of the potassium salt of 2‑methylproline‑2‑carboxylate was observed when the reaction mixture temperature dropped below 5 °C during aqueous work‑up. This necessitated installation of a jacketed reactor with propylene glycol circulation to maintain a minimum working temperature of 10 °C and avoid line blockage.
| Parameter | (R)-Boc-2‑methylproline | (S)-Boc-2‑methylproline | Boc-L-proline | (R)-Fmoc-2‑methylproline |
|---|---|---|---|---|
| CAS RN | 1034707-13-1 | 1034707-12-0 | 15761-39-4 | 1270587-34-9 |
| Molecular weight | 229.27 | 229.27 | 215.25 | 353.40 |
| Purity specification | ≥98.5% (HPLC) | ≥98.5% (HPLC) | ≥99.0% (HPLC) | ≥97.0% (HPLC) |
| Enantiomeric excess | ≥99.0% | ≥99.0% | N/A (achiral) | ≥99.0% |
| Water content limit | ≤0.5% | ≤0.5% | ≤0.5% | ≤1.0% |
| Racemization risk during coupling | None (quaternary Cα) | None (quaternary Cα) | 1–3% per cycle | None (quaternary Cα) |
| Deprotection method | 95% TFA, 30–60 min | Identical | Identical | 20% piperidine/DMF, 2 × 5 min |
| Recommended coupling time | 2 × 15 min (microwave) | Identical | 5 min (microwave) | 10 min (microwave) |
Industrial users report that switching from Boc‑proline to Boc‑2‑methylproline in the assembly of a CXCR4 antagonist heptapeptide eliminated a late‑stage D‑proline contaminant that previously required preparative HPLC purification. The increased steric demand necessitated raising the synthesis temperature from 75 °C to 90 °C during the coupling step, which was accommodated by a CEM Liberty PRIME instrument with a calibrated internal thermocouple. This alteration exemplifies the precise equipment‑dependent adjustments that the quaternary amino acid imposes.
| Regulation/Standard | Status | Reference |
|---|---|---|
| US Toxic Substances Control Act (TSCA) | Inventory‑listed | Section 8(b) |
| REACH (EU) Regulation (EC) No 1907/2006 | Pre‑registration completed for 1–10 t/a band | Article 28 |
| Global Harmonized System (GHS) classification | Skin Irrit. 2, Eye Irrit. 2A, STOT SE 3 | EC 1272/2008 |
| Controlled Substance Act (US DEA) | Not scheduled | 21 CFR 1308.11–1308.15 |
| FDA 21 CFR Food Additive Status | Not approved as direct food additive | 21 CFR 170.3 |
| Dual‑use export control | Not listed | EU 2021/821 |
When large‑scale peptide API production exceeds 500 g of incorporated (R)‑2‑methylproline residue, compatibility with high‑throughput SPPS reactors (e.g., Syro Wave™ parallel synthesizer) is critically dependent on adequate pre‑drying of the amino acid. In one campaign, residual moisture of 1.2% from an opened container led to Boc deprotection during the coupling step and generated a truncated peptide sequence that co‑eluted with the target peptide on preparative C18 columns. The resulting batch reprocessing consumed 6 hours of preparative HPLC time and increased solvent consumption by 2.4‑fold. Subsequent implementation of a nitrogen‑purge loading station with integrated Karl Fischer monitoring brought moisture below 0.2% and eliminated the failure mode.
The compound is not classified as flammable, but thermal decomposition above 150 °C releases carbon dioxide and isobutylene; ventilation must be adequate during hot‑plate drying protocols. Waste disposal follows standard procedures for non‑halogenated organic laboratory chemicals, with incineration preferred over hydrolytic treatment to avoid liberation of the biologically active 2‑methylproline core into wastewater streams.
