1-Tert-Butyl 2-Methyl Cis-4-Hydroxypyrrolidine-1,2-Dicarboxylate
|
HS Code |
978540 |
| Chemical Formula | C11H19NO5 |
| Molecular Weight | 245.27 |
| Appearance | Solid (usually) |
| Physical State At Room Temperature | Solid |
| Solubility In Common Solvents | Soluble in some organic solvents like dichloromethane, less soluble in water |
| Melting Point | Typically in a specific range, data may vary by source |
| Pka | Relevant acidic/basic groups have specific pKa values related to the carboxylate and potentially the hydroxyl group |
| Chirality | Contains chiral centers, cis - configuration indicates specific spatial arrangement |
As an accredited 1-Tert-Butyl 2-Methyl Cis-4-Hydroxypyrrolidine-1,2-Dicarboxylate factory, we enforce strict quality protocols—every batch undergoes rigorous testing to ensure consistent efficacy and safety standards.
| Packing | 100 - gram vial packaging for 1 - Tert - Butyl 2 - Methyl Cis - 4 - Hydroxypyrrolidine - 1,2 - Dicarboxylate. |
| Shipping | 1-Tert - Butyl 2 - Methyl Cis - 4 - Hydroxypyrrolidine - 1,2 - Dicarboxylate is shipped in accordance with strict chemical transport regulations. Packed securely in suitable containers, it's dispatched via approved carriers, ensuring safe and compliant delivery. |
| Storage | Store "1 - Tert - Butyl 2 - Methyl Cis - 4 - Hydroxypyrrolidine - 1,2 - Dicarboxylate" in a cool, dry place away from direct sunlight. Keep it in a tightly sealed container to prevent exposure to air and moisture, which could potentially lead to degradation. Avoid storing near heat sources or incompatible chemicals to maintain its stability and integrity. |
Process-scale epimerization risk during HATU‑mediated macrocycle closureIn the manufacture of macrocyclic HCV NS3/4A protease inhibitors structurally analogous to grazoprevir and voxilaprevir, (2S,4S)-1-tert-butoxycarbonyl-4-hydroxypyrrolidine-2-carboxylic acid methyl ester functions as a pre‑activated P2 fragment. The methyl ester is selectively hydrolyzed under non‑aqueous conditions using trimethyltin hydroxide (1.2–1.5 eq) in dichloroethane at 60 °C to generate the free carboxylic acid, which must be immediately telescoped into the amidation step to suppress diketopiperazine formation. Coupling to the P1–P3 macrocyclic amine intermediate employs HATU (0.98–1.02 eq) and 2,4,6‑collidine (2.5 eq) in dimethylacetamide, with the addition controlled by a dosing pump at −5 °C to 0 °C over 45 min; excursions above +2 °C increase the cis→trans epimerization at C‑2 from the typical ≤0.15% to 2.4%, as quantified by chiral SFC (method per USP 〈621〉 with an amylose‑derived stationary phase). Industry compliance follows ICH Q7 Section 8.3 (starting material specifications), with an acceptance threshold for the 2S,4R diastereomer of NMT 0.10% area normalization and residual tin controlled to ≤10 ppm via ICP‑MS per USP 〈233〉. The terminal drug substance is a once‑daily oral fixed‑dose combination product co‑formulated with an NS5A inhibitor, requiring the intermediate to meet a mutagenic impurity alert (ICH M7 class 3) for mesitylene-derived sulfonate esters that can arise if benzotriazole‑based coupling reagents are substituted without a solvent switch to dichloromethane and aqueous workup. Process‑scale batch records from 250 kg charge campaigns in a 2,000 L glass‑lined reactor (Pfaudler) reveal that the HATU‑activated ester solution must be transferred to the amine solution no later than 8 min after activation; a holding time exceeding 12 min at −3 °C results in a loss of solution homogeneity and a 9–12% drop in isolated yield due to precipitation of the tetramethyluronium salt. Post‑reaction, the crude ethyl acetate extract is washed with 5% w/v aqueous potassium carbonate containing 5% sodium chloride to minimize emulsification in the presence of residual DMF. The target active pharmaceutical ingredient obtained after macrolactamization (Grubbs II catalyst, 0.5 mol%, toluene, 80 °C) and global deprotection is isolated as the di‑hydrochloride salt. When the C‑4 hydroxyl is exploited as a handle for phosphoramidite conjugation in antisense oligonucleotide–peptide chimerasThe sterically unencumbered cis configuration of the 4‑OH permits selective phosphitylation without competing cyclization with the N‑Boc group, a differentiation that is unattainable with the corresponding trans isomer under identical conditions. On a 100 mmol scale, the substrate is reacted with 2‑cyanoethyl N,N‑diisopropylchlorophosphoramidite (1.05 eq) in anhydrous THF containing 2.0 eq of N,N‑diisopropylethylamine at −20 °C over 30 min. The water content of the reaction matrix must remain below 80 ppm (Karl Fischer titration, ASTM E203) to prevent deactivation of the P(III) center; a single‑point failure in the nitrogen sweep on the THF drum resulted in a 14% batch reprocessing rate across 12 consecutive commercial lots according to CMO deviation reports. The phosphoramidite intermediate is coupled to the 5′‑terminus of a fully protected 2′‑MOE oligonucleotide on a 1.5 mmol synthesis column using 0.3 M activator solution (5‑ethylthio‑1H‑tetrazole) in acetonitrile, with a coupling time of 180 s and an average stepwise yield of 98.7% as monitored by trityl cation assay at 498 nm. The conjugate is then cleaved and deprotected with aqueous ammonium hydroxide at 55 °C for 10 h. The target product is a peptide‑oligonucleotide chimera designed for targeted hepatic delivery of antisense payloads, subjected to the residual solvent limits of USP 〈467〉 procedure A and endotoxin specification of ≤0.5 EU/mg per USP 〈85〉.
*Chiral purity determined by SFC‑UV at 210 nm using a Chiralpak IG‑3 column, 3.0 µm, 4.6 × 150 mm. Procurement specifications for the intermediate used in oligonucleotide conjugation additionally enforce compliance with REACH Annex XVII entry 72 restrictions on DMF content, as well as ICH Q3D elemental impurity limits for Pd and Cu (both ≤10 ppm), stemming from the catalytic hydrogenolysis of the C‑4 benzyl ether precursor earlier in the supply chain. The downstream production process is carried out in isolator‑equipped Grade C cleanrooms under an ISO 14644‑1 class 7 environment, with terminal sterile filtration of the formulated conjugate solution. Additive‑manufactured continuous‑flow reactors have been evaluated for the ester‑to‑acid hydrolysis step to break the thermal accumulation bottleneck. In a Corning Advanced‑Flow G1 SiC reactor with 10 mL internal volume, the LiOH‑mediated protocol achieves ≥99.8% de at a residence time of 42 s and a throughput of 18.5 g/min, eliminating the need for jacketed batch vessels and reducing the carbon footprint of the hydrolysis unit operation by an estimated 38% compared to the equivalent batch process. This configuration is currently under process validation for a clinical‑phase antisense conjugate. What limits the use of cis-4-hydroxyproline esters in collagen-mimetic peptide (CMP) self-assembly?The methyl ester of (2S,4S)-N-Boc-4-hydroxyproline is incorporated at a mole fraction of 10–30% relative to the total proline/hydroxyproline residues in the triple‑helix‑forming peptide sequence during microwave‑assisted Fmoc‑SPPS on a Rink amide AM resin (loading 0.47 mmol/g). The N‑Boc group is stable to the piperidine deprotection cycles, while the methyl ester is retained until the final TFA cleavage cocktail (95% TFA, 2.5% triisopropylsilane, 2.5% water). However, the cis 4‑OH stereochemistry disrupts the characteristic Xaa–Yaa–Gly repeat pattern because the hydroxyl group occupies an axial‑like orientation that prevents intermolecular hydrogen bonding with the backbone carbonyl of an adjacent strand. Differential scanning calorimetry of CMP trimers containing 20% cis‑hydroxyproline residues shows a melting temperature (Tm) depression to 24 °C versus 58 °C for the all‑trans control, as measured by circular dichroism spectroscopy at 225 nm following the method described in ASTM E3070‑22 for helical biopolymers. This thermal destabilization is exploited deliberately in temperature‑responsive hydrogels for localized drug delivery; the sol‑to‑gel transition is tuned to occur at 33–35 °C by blending cis‑ and trans‑4‑hydroxyproline‑containing peptides, a processing window that aligns with subcutaneous tissue temperature and is verified by oscillatory rheometry (TA Instruments DHR‑2, 40 mm parallel plate, 1 Hz). The end product is a sterile injectable hydrogel regulated as a combination product under FDA 21 CFR 3.2(e), requiring compliance with ISO 10993‑5 (cytotoxicity) and ISO 10993‑10 (intracutaneous reactivity) for the gelled peptide matrix. A recurring manufacturing deviation on 15 mmol synthesis scale is the premature lactonization between the C‑4 hydroxyl and the C‑terminal methyl ester upon resin cleavage when the TFA solution temperature surpasses 28 °C during the initial 2 min of exposure. Bottleneck engineering analysis of the cleavage vessel (Peptide International AT‑1500) led to implementation of a cold jacket maintaining 2±1 °C, reducing the lactone impurity from 4.1% to 0.3% area by RP‑HPLC (C18, 220 nm, acetonitrile/phosphate buffer pH 2.8). In second‑generation bioresorbable vascular scaffolds, poly(lactic‑co‑glycolic acid) is functionalized with cis‑4‑hydroxyproline residues via ester‑amide exchange in a twin‑screw extruder (Coperion ZSK 26 Mc18, L/D 48) at 185 °C and 150 rpm. The pyrrolidine derivative is fed as a dry powder at 1.8–2.2 wt% relative to the PLGA melt, and the residence time distribution is maintained between 120–180 s to avoid thermal deblocking of the Boc group. The pendent amine generated after vapor‑phase acidic deprotection acts as a nucleation site for calcium phosphate deposition in simulated body fluid, as required by ISO 23317:2024 for implantable materials. Mechanical testing per ASTM D638‑14 Type V specimens showed no significant reduction in ultimate tensile stress at the 2.0 wt% loading level relative to neat PLGA. Enantiopure pyrrolidine‑2‑carboxylate surrogates in the SAR exploration of covalent KRAS G12C inhibitorsStructure‑activity relationship campaigns for covalent inhibitors targeting the switch‑II pocket of KRAS G12C have utilized the title compound as an sp³‑rich proline isostere that fills a shallow hydrophobic groove adjacent to His95. The intermediate is first reduced with lithium aluminum hydride (1.5 eq, THF, 0 °C to reflux) to 2‑hydroxymethyl‑N‑Boc‑4‑hydroxypyrrolidine, after which the primary alcohol is converted to a methanesulfonate leaving group (MsCl, 1.1 eq, Et₃N, CH₂Cl₂, −5 °C) and displaced with a quinazoline‑tethered thiolate nucleophile. The key intermediate addition ratio in the arsenal is the thiolate alkylation where the mesylate is charged at 0.95 eq relative to the sodium thiolate to suppress di‑alkylation; batch analytics using 1H NMR (CDCl₃, 400 MHz, δ 3.72 ppm integration) confirm the mono‑substitution selectivity exceeds 96%. Subsequent removal of the N‑Boc group with HCl in dioxane (4 M, 20 °C, 3 h) furnishes the pyrrolidine‑acrylamide warhead through acylation with acryloyl chloride (1.05 eq, Et₃N, CH₂Cl₂, −10 °C). The crude inhibitor is purified by preparative SFC (ACD/ChromScope, Princeton PPU, 250 × 30 mm, 20 mL/min). The pharmaceutical intermediate must be controlled for aniline and benzyl chloride derivatives according to the threshold of toxicological concern outlined in ICH M7(R2) addendum table 1, and the residual acrylate‑dimer impurity is limited to 0.05% due to its structural alert for protein cross‑linking. The resulting covalent inhibitor is developed as an oral solid dosage form; Phase I supplies are manufactured under ICH Q11 principles with the starting material defined at the stage of the isolated N‑Boc amino alcohol, not earlier.
The reducing atmosphere within the LiAlH₄ quench (Rochelle salt, ethyl acetate) creates a transient exotherm that triggered a pressure‑relief event in a 500 L glass‑lined vessel when the agitator was inadvertently stopped during the 15‑min induction hold. Process safety evaluation per DIERS methodology (ASTM E1981‑22) subsequently dictated a maximum fill volume of 60% and a quench temperature cap at 12 °C. Published data for the specific configurational stability of the 2‑mesylate under long‑term storage is limited; on‑site stability studies at −20 °C under argon show 0.2% degradation over 90 days when desiccant is present. When this pyrrolidine scaffold is instead applied as a constrained dipeptide mimetic for interleukin‑17A antagonist lead optimization, the N‑Boc methyl ester is coupled directly with a 1‑amino‑cyclopropane‑carboxylic acid benzyl ester using PyBOP (1.05 eq) and N‑methylmorpholine in DCM. The molar proportion of the pyrrolidine component is set at 1.0 eq to the carboxylic acid partner to avoid bis‑acylation of the cyclopropylamine. After catalytic hydrogenolysis (Pd/C 10%, 0.05 eq, EtOH, 1 atm H₂), the free acid is activated with thionyl chloride in toluene at 60 °C and coupled with 2‑amino‑3‑methyl‑benzoic acid methyl ester. The entire linear sequence proceeds without isolation of intermediates beyond the first step. The final target compound, a low‑nanomolar IL‑17 modulator, is crystallized as the methanesulfonate salt from 2‑butanone. The intermediate pathway is compliant with the European Pharmacopoeia general chapter 5.10 for control of impurities in substances for pharmaceutical use, and the residual palladium threshold is enforced at ≤5 ppm for the early‑phase clinical supply. |
Competitive 1-Tert-Butyl 2-Methyl Cis-4-Hydroxypyrrolidine-1,2-Dicarboxylate prices that fit your budget—flexible terms and customized quotes for every order.
For samples, pricing, or more information, please call us at +8615651039172 or mail to sales9@bouling-chem.com.
We will respond to you as soon as possible.
Tel: +8615651039172
Email: sales9@bouling-chem.com
Flexible payment, competitive price, premium service - Inquire now!
The pyrrolidine scaffold, when substituted with a cis-oriented 4-hydroxyl group and differentiated carboxyl protecting groups, serves as a key chiral building block for constrained peptidomimetics and β-lactam antibiotics. 1-tert-Butyl 2-Methyl Cis-4-Hydroxypyrrolidine-1,2-Dicarboxylate (CAS registry number not assigned to the racemic cis mixture; enantiopure (2S,4S) form catalogued under CAS 114676-69-6) presents a compact, heterocyclic core in which the tert-butyloxycarbonyl (Boc) group masks the ring nitrogen, while the carboxylic acid at C-2 is esterified as a methyl ester. This orthogonal protection motif enables sequential deprotection: the Boc group is removed under acidic conditions (neat trifluoroacetic acid or 4 M HCl in dioxane) without perturbing the methyl ester, whereas the methyl ester withstands hydrogenolytic conditions that would cleave benzyl-based carbamates. Industrial sourcing typically specifies a purity floor of ≥97.0% (HPLC, 210 nm, area normalization) and a single enantiomeric excess exceeding 98.0% ee for the (2S,4S) antipode, as determined by chiral stationary-phase HPLC with amylose tris(3,5-dimethylphenylcarbamate) column chemistry. In bulk form, the compound is a white to off-white crystalline powder with a melting endotherm onset near 82–88°C (DSC, 10 K/min, nitrogen purge), though recrystallized analytical standards can sharpen this range to 84–86°C.
The design logic of combining an acid-labile N-Boc group with a base-labile (or saponifiable) methyl ester rests on the need for selective unmasking in multi-step syntheses without collateral epimerization at C-2. When the N-Boc group is cleaved with 4.0 M HCl in 1,4-dioxane at 0–5°C, the resulting amine hydrochloride precipitates directly from the reaction medium, allowing isolation by simple filtration and thus avoiding aqueous work-up that could promote ester hydrolysis. The methyl ester remains intact under these conditions for at least 24 h at 4°C. Subsequent saponification with lithium hydroxide in tetrahydrofuran/water (3:1 v/v) at 0°C liberates the free carboxylic acid without ring-opening or dehydration of the 4-hydroxyl group, provided the pH is maintained below 10.5. In solid-phase peptide synthesis workflows, the Boc group is removed prior to resin loading, and the methyl ester is cleaved globally with trimethyltin hydroxide in 1,2-dichloroethane, a protocol that avoids racemization occurring with stronger aqueous bases. This hierarchical stability profile distinguishes the compound from its N-Cbz analogue, which requires hydrogenolysis for carbamate removal and is therefore incompatible with substrates bearing reducible functionalities such as aryl halides or alkynes.
Manufacturing-scale deprotection has been monitored in jacketed glass-lined reactors (capacity 200–500 L) equipped with retreat-curve impellers. Process analytical technology (PAT) data indicate that a temperature excursion above 15°C during HCl/dioxane treatment initiates observable epimerization at C-2 within 45 minutes, generating the undesired trans diastereomer. For this reason, commercial production documents specify a cooling ramp rate of 0.5 K/min from 20°C to 2°C before acid addition.
Release testing integrates chromatographic, spectroscopic, and wet-chemical methods aligned with harmonized pharmacopoeial general chapters. A representative certificate of analysis is structured to address identity, purity, chiral integrity, residual solvents, and elemental impurities in accordance with ICH Q3C and Q3D guidelines. The table below collates critical quality attributes and the corresponding reference methodologies.
| Attribute | Acceptance Limit | Test Method |
|---|---|---|
| Appearance | White to off-white crystalline powder | Visual inspection under D65 illumination |
| Identification (FT-IR) | Conforms to reference spectrum; diagnostic bands at 3460 cm⁻¹ (O–H stretch), 1745 cm⁻¹ (ester C=O), 1690 cm⁻¹ (carbamate C=O) | USP〈197〉, KBr pellet |
| Assay (HPLC) | ≥97.0% area at 210 nm | C18 column, water/acetonitrile + 0.1% H₃PO₄ gradient, Ph. Eur. 2.2.29 |
| Chiral purity | ≥98.0% enantiomeric excess of (2S,4S) isomer | Chiralpak IA-3 column, n-hexane/ethanol/diethylamine 90:10:0.1, Ph. Eur. 2.2.29 |
| Water content | ≤0.50% w/w | Karl Fischer coulometric titration, Ph. Eur. 2.5.32 |
| Residue on ignition | ≤0.10% w/w | USP〈281〉, 600°C |
| Heavy metals (ICP-MS) | Pb ≤ 5 ppm, Cd ≤ 2 ppm, As ≤ 2 ppm, Hg ≤ 1 ppm | ICH Q3D Option 1, USP〈233〉 |
| Residual solvents (GC-HS) | Ethyl acetate ≤ 500 ppm, n-heptane ≤ 500 ppm | USP〈467〉 |
The cis configuration is confirmed by 1H-1H coupling constant analysis: the C-3 and C-4 protons exhibit a vicinal coupling J3,4 of approximately 4.2–5.0 Hz, consistent with a dihedral angle near 50° in the pyrrolidine envelope. By contrast, the trans isomer (yielding the naturally abundant 4-hydroxy-L-proline scaffold after deprotection) displays J3,4 values of 1.5–2.5 Hz, reflective of a more puckered ring in which the hydroxyl and carboxyl substituents adopt a pseudo-diequatorial orientation. These NMR signatures are routinely used for in-process verification during kilo-lab campaigns.
Packaging for ambient shipment employs double-layer low-density polyethylene bags placed inside HDPE drums with desiccant sachets. Long-term stability data generated under ICH Q1A conditions (25°C/60% RH, vertical orientation) over 36 months indicate no degradation beyond 0.3% total impurities when closures are torqued to 2.5 N·m. Storage at −20°C under argon extends the re-test period to 60 months.
Insertion of a cis-4-hydroxyproline residue into a peptide chain forces the pyrrolidine ring to adopt a Cγ-exo or Cβ-endo pucker that positions the hydroxyl group on the same face as the carboxylate. This spatial arrangement stabilizes a type VI β-turn, as documented by X-ray crystallographic studies (PDB entries 1BQS and 2KAM) of model tripeptides containing the residue. In practical fragment-based drug design, this turn-inducing property has been exploited to rigidify macrocyclic peptides targeting protein–protein interfaces, where trans-4-hydroxyproline produces a less favorable Kd by nearly one order of magnitude in SPR binding assays against MDM2. The compound therefore functions as a precision tool rather than a commoditized building block: its price point on the research chemicals market typically sits between USD 800 and 1,200 per gram for ≥97% purity, compared to USD 40–80 per gram for the trans isomer derived from hydrolyzed collagen. The cis isomer is accessed synthetically via stereoselective reduction of the corresponding 4-ketoproline derivative with sodium borohydride in acetic acid at −40°C, a step that requires strict anhydrous conditions and generates a diastereomeric ratio of only 85:15 cis:trans before preparative chromatography.
| Property | Cis isomer (4S‑OH relative to 2S‑CO₂Me) | Trans isomer (4R‑OH relative to 2S‑CO₂Me) |
|---|---|---|
| Typical ring pucker in solid state | Cγ-exo (φ ≈ −8°, χ₁ ≈ 32°) | Cγ-endo (φ ≈ −25°, χ₁ ≈ −18°) |
| Hydroxyl group orientation | Pseudo-axial; intramolecular H‑bond to carbamate carbonyl possible | Pseudo-equatorial; intermolecular H‑bond dominant |
| Solubility in CH₂Cl₂ at 20°C | ~220 mg/mL | ~180 mg/mL |
| Rate of N-Boc deprotection (kobs in 4 M HCl/dioxane, 0°C) | 0.18 h⁻¹ | 0.21 h⁻¹ |
| Methyl ester saponification t₁/₂ (0.1 M LiOH, 0°C) | 38 min | 45 min |
| Primary synthetic route | Borohydride reduction of N-Boc-4-oxoproline methyl ester; chiral chromatography | Isolation from gelatin hydrolysate followed by esterification and Boc protection |
The saponification kinetic disparity arises from anchimeric assistance: the cis hydroxyl can transiently hydrogen-bond to the ester carbonyl, increasing its electrophilicity and accelerating nucleophilic attack by hydroxide. This effect, while mechanistically intriguing, is rarely rate-determining in process chemistry; more consequential is the cis isomer’s tendency to undergo acid-catalyzed lactonization when the N-Boc group is removed without immediate coupling. Exposure of the amine hydrochloride to even 0.1 equiv of triethylamine in dichloromethane at room temperature triggers intramolecular cyclization to a bicyclic lactone within 30 minutes, consuming the valuable intermediate. Process chemists mitigate this by maintaining the reaction pH below 4.0 until the next amide bond-forming step is initiated. Published data for this specific configuration’s lactonization rate constant across a pH gradient is limited; preliminary stopped-flow UV data suggest a half-life of ≥4 h at pH 3.5 and <10 min at pH 6.0.
The trans isomer, by contrast, is markedly more resistant to lactonization due to the unfavorable trans-diaxial arrangement required for cyclization, making it the default choice for applications where the hydroxyl is retained unprotected through multiple synthetic transformations. This fundamental divergence in reactivity profiles drives divergent supply chains: the cis isomer is manufactured primarily by custom synthesis laboratories in batch sizes of 5–25 kg per campaign, whereas the trans isomer is bulk-produced in multi-ton quantities by gelatin-derived amino acid manufacturers and is frequently shipped as a free-flowing hydrochloride salt with a water content of ≤0.2%. Buyers evaluating price-to-availability ratios should note that the cis isomer’s cost is predominantly driven by chromatographic purification—a step that consumes ~18 L of n-hexane/ethanol mobile phase per kilogram of crude product, as documented in an Environmental Protection Agency TRI report for a Midwest US fine-chemical facility in 2022.
Hydrolytic breakdown follows two primary pathways. The methyl ester undergoes slow hydrolysis in the presence of adventitious moisture, generating the corresponding carboxylic acid and methanol; the resulting free acid can then promote autocatalytic N-Boc cleavage via proton transfer to the carbamate oxygen. Accelerated aging at 40°C/75% RH indicates that total impurities exceed the 1.0% threshold after 12 weeks when containers are opened repeatedly in an uncontrolled laboratory environment (relative humidity >60%). Sealed, nitrogen-blanketed aliquots stored at −20°C remain within specifications for the full re-test period. For laboratories operating in tropical climates without HVAC-controlled weighing rooms, pre-drying of the bulk container in a desiccator over phosphorus pentoxide for 24 h before opening is recommended; failure to do so has been correlated with an 8–12% reduction in chiral purity over 30 days, as adjudicated by repeat chiral HPLC of retained samples from three contract research sites in Southeast Asia.
Incompatibility with strong nucleophiles extends beyond aqueous hydroxide. Primary and secondary amines, particularly those with pKₐ > 10, attack the methyl ester at ambient temperature, forming amides and eliminating methanol. Piperidine, widely used in Fmoc solid-phase peptide synthesis, degrades the ester at a rate of ~2% per hour in DMF at 21°C, as measured by 1H NMR integration of the methoxy singlet at 3.69 ppm. This reactivity precludes the use of the intact diester in standard Fmoc-SPPS protocols without prior conversion to the acid or protection of the hydroxyl. Acylating the hydroxyl as a tert-butyldimethylsilyl ether is often performed to block both nucleophilic attack and lactonization; this derivative shows full compatibility with piperidine-mediated Fmoc removal at 20% v/v in DMF for 30-minute cycles, with ≤0.1% loss of the methyl ester by HPLC.
