5-(Z)-(5-Fluoro-2-Oxoindolin-3-Ylidene)Methyl)-2,4-Dimethyl-1H-Pyrrole-3-Carboxylic Acid

Product Name 5-(Z)-(5-Fluoro-2-Oxoindolin-3-Ylidene)Methyl)-2,4-Dimethyl-1H-Pyrrole-3-Carboxylic Acid
Alias JK-312
Einecs 821-617-2
Mininmum Order 1mg
Factory Site West Ujimqin Banner, Xilingol League, Inner Mongolia, China
Price Inquiry sales9@bouling-chem.com
Manufacturer Bouling Chemical Co., Limited
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In terms of specification, 5-(Z)-(5-Fluoro-2-Oxoindolin-3-Ylidene)Methyl)-2,4-Dimethyl-1H-Pyrrole-3-Carboxylic Acid is supplied with HPLC purity and Z-isomer content, making it suitable for pharmaceutical intermediate synthesis.

Specifications

HS Code	378063
Chemical Formula	C16H13FNO4
Molecular Weight	301.28
Iupac Name	5-[(Z)-(5-fluoro-2-oxoindolin-3-ylidene)methyl]-2,4-dimethyl-1H-pyrrole-3-carboxylic acid

As an accredited 5-(Z)-(5-Fluoro-2-Oxoindolin-3-Ylidene)Methyl)-2,4-Dimethyl-1H-Pyrrole-3-Carboxylic Acid factory, we enforce strict quality protocols—every batch undergoes rigorous testing to ensure consistent efficacy and safety standards.

Packing & Storage

Packing	100g of 5-(Z)-(5 - Fluoro - 2 - Oxoindolin - 3 - Ylidene)Methyl)-2,4 - Dimethyl - 1H - Pyrrole - 3 - Carboxylic Acid in sealed container.
Shipping	The chemical "5-(Z)-(5 - Fluoro - 2 - Oxoindolin - 3 - Ylidene)Methyl)-2,4 - Dimethyl - 1H - Pyrrole - 3 - Carboxylic Acid" will be shipped in specialized, sealed containers, following strict chemical transport regulations to ensure safety during transit.
Storage	Store "5-(Z)-(5 - Fluoro - 2 - Oxoindolin - 3 - Ylidene)Methyl)-2,4 - Dimethyl - 1H - Pyrrole - 3 - Carboxylic Acid" in a cool, dry place, away from heat sources and direct sunlight. Keep it in a tightly - sealed container to prevent moisture absorption and potential reactions with air components. Avoid storing near incompatible chemicals.

Application of 5-(Z)-(5-Fluoro-2-Oxoindolin-3-Ylidene)Methyl)-2,4-Dimethyl-1H-Pyrrole-3-Carboxylic Acid

In multi-kilogram production campaigns for sunitinib malate conducted under full cGMP conditions aligned with ICH Q7 (Part II, Active Pharmaceutical Ingredients) and starting material designation per ICH Q11 Section 5, the 5-[(Z)-(5-fluoro-2-oxoindolin-3-ylidene)methyl]-2,4-dimethyl-1H-pyrrole-3-carboxylic acid intermediate is charged as the key indolinone-pyrrole building block immediately prior to the pivotal amide bond-forming step. Because this intermediate bears both the critical Z-configured exocyclic double bond and the free carboxylic acid handle, its purity, residual solvent profile, and isomeric integrity directly influence the yield, impurity fate, and subsequent regulatory acceptance of the final drug substance. The stoichiometric molar feed ratio of the intermediate to N,N-diethylethylenediamine (side chain) is maintained between 1.0:1.10 and 1.0:1.25 to drive the amidation to completion after in situ activation of the carboxyl group with 1,1′-carbonyldiimidazole (CDI) in anhydrous dimethylformamide at a jacket temperature of 0–5°C; this narrow excess ensures that the residual primary amine is manageable in the subsequent aqueous work-up while preventing the formation of dimeric side products that are otherwise observed when the stoichiometry deviates beyond 1.0:1.30. The downstream manufacturing sequence proceeds with quenching into purified water, controlled crystallization of the sunitinib free base from acetone/water mixtures, reslurry in 2-propanol to purge unreacted indolinone and the corresponding E-isomer, and final salt formation with L-malic acid in ethanol to furnish sunitinib malate conforming to the specifications of USP and Ph.Eur. monographs. Residual solvents are controlled according to ICH Q3C(R8): dimethylformamide (Class 2) is limited to ≤ 880 ppm, acetone (Class 3) to ≤ 5000 ppm, and 2-propanol to ≤ 5000 ppm. On production-scale equipment—typically glass-lined reactors of 4000–6300 L capacity equipped with amber-lighting filtering systems and completely shrouded sight glasses—the entire batch process is executed under strict exclusion of light below 520 nm because the Z-isomer undergoes facile photoisomerization to the biologically inactive E-configuration; in full-scale campaigns, a photostability audit of the supply chain is imposed, and any lot exhibiting a Z/E ratio below 99.5:0.5 by HPLC area is diverted to a re-crystallization loop prior to amidation. The factory acceptance criterion for the intermediate mandates a purity of ≥ 99.5% (sum of Z-isomer), single unknown impurity ≤ 0.10%, and total impurities ≤ 0.50%, all quantified against a certified reference standard by the in-house quality control unit using a validated C18 reverse-phase HPLC method with UV detection at 268 nm. The terminal dosage form manufactured from the resulting API is sunitinib malate hard gelatin capsules in strengths of 12.5 mg, 25 mg, and 50 mg.

What Limits the Reporting Threshold for the Z-Isomer in Chronic-Use Oncology Formulations?

During the development and commercial lifecycle of sunitinib malate oral capsules, the 5-fluoroxindole intermediate—and specifically its Z-geometric configuration—serves as a primary impurity marker in stability-indicating analytical methods mandated by ICH Q3B(R2) for degradation products in finished dosage forms. Because the maximum recommended daily dose of sunitinib is 50 mg, the reporting threshold for any specified degradation product is set at 0.2% of the labeled strength, and the identification threshold is 0.5%; the identification quantity corresponds to a maximum of 0.25 mg per daily intake, a level that triggers retrieval of retention time and UV spectral identity against a reference standard of the Z-intermediate. For method validation, the reference standard is spiked into a placebo blend consisting of mannitol, crospovidone, colloidal silicon dioxide, and magnesium stearate at three concentration levels: 0.10%, 0.20%, and 0.50% (w/w relative to the 12.5 mg capsule content), and the recovery study performed on a fully automated Zanasi LZ64 encapsulation line that doses the powder mixture into size 3 hard gelatin shells. The quantitative HPLC method employs a 150 × 4.6 mm, 3.5 µm C18 column maintained at 30°C, a mobile phase of phosphate buffer (pH 3.0) and acetonitrile in gradient mode at a flow rate of 1.0 mL/min, and UV detection at 268 nm to achieve baseline resolution between the Z-isomer and the sunitinib peak; the system suitability requirement demands a resolution factor of ≥ 2.0 between the E- and Z-isomers, with the Z-isomer eluting as the later peak. Across forty-five independent determinations at the three spiking levels, the overall percent recovery for the intermediate must fall within 95.0%–105.0% with a relative standard deviation not exceeding 3.0%—a window established through collaborative trial data run at contract manufacturing organizations operating under EU GMP Annex 11 computerized systems. Terminal finished product types are the sunitinib malate hard capsules in immediate-release presentations; any batch in accelerated stability testing (40°C ± 2°C / 75% RH ± 5% RH) that shows a Z-isomer increase above the 0.5% identification threshold triggers an out-of-specification investigation per the quality agreement and may lead to accelerated photoprotective packaging studies.

In forced degradation chambers set up under ICH Q1B Option 2 conditions—exposing milled sunitinib malate powder and spiked placebo blends to 1.2 million lux·hours of visible light and integrated near-UV energy of 200 W·h/m²—the 5-fluorooxindole-3-ylidene intermediate rapidly photoisomerizes via a triplet excited state, producing an equilibrium mixture in which the E-isomer area percent reaches 12–18% by HPLC after 24 hours of exposure in dilute methanol solution; this photolability makes the pure Z-intermediate reference standard an indispensable tool for constructing system suitability mixtures for photodegradation method validations. The reference standard is prepared by dissolving the intermediate in dimethyl sulfoxide at a concentration of 0.5 mg/mL, spiking the solution with a pre-irradiated batch containing 0.5% (v/v) of the E-isomer marker, and dispensing aliquots into amber USP Type I glass vials that are sealed under argon and stored at -20°C ± 5°C to suppress further isomerization. Certification of the standard involves quantitative ¹H NMR spectroscopy (qNMR) following ISO/IEC 17025:2017 general requirements and ISO Guide 35:2017 for reference material characterization, using dimethyl sulfone as an internal calibrant and acquiring spectra on a 600 MHz spectrometer with a 30-second relaxation delay to ensure full proton relaxation. The certified value of the Z-isomer content is assigned an expanded uncertainty of ± 0.4% (k = 2). Downstream proprietary certified reference material production is conducted in a cleanroom environment (ISO Class 7) and the final standards are delivered as 100 mg units in crimp-sealed amber vials with a lot-specific Certificate of Analysis compliant with ISO 17034:2016. The end-use finished product is not a pharmaceutical dosage form but rather an analytical reference standard used by quality control laboratories, contract analytical service providers, and pharmacopoeial standards verification programs to calibrate HPLC-UV and LC-MS/MS methods intended for genotoxic impurity and degradation product profiling in sunitinib formulations.

When the Indolinone Intermediate Demonstrates Nitrosamine Absence, ANDA DMF Acceptability Accelerates

A nitrosamine-free status for the 5-fluoroxindole-3-ylidene intermediate has evolved into a pre-market gatekeeping requirement that directly determines the approvability of abbreviated new drug applications (ANDAs) referencing sunitinib malate. Even though the pyrrole ring in the intermediate possesses a secondary amine that could, in principle, react with nitrosating agents, well-controlled manufacturing routes eliminate any plausible source of exogenous nitrite, and the absence of N-nitrosamine impurities must be demonstrated in the submission’s Drug Master File (DMF) conducted according to ICH M7(R2) and the FDA Guidance for Industry: Control of Nitrosamine Impurities in Human Drugs (revised February 2021). The acceptable intake for any single N-nitrosamine impurity is provisionally capped at 26.5 ng/day based on the carcinogenic potency categorization approach for a lifetime exposure cancer risk of 1 in 100,000; for a maximum daily dose of 50 mg sunitinib, this translates to a concentration limit of 530 ppb (or 0.53 ppm) in the drug substance. To provide an adequate safety margin, the intermediate supplier’s specification imposes a tighter limit: any N-nitrosamine detected by a validated LC-MS/MS method using atmospheric pressure chemical ionization must be below the 0.03 ppm (w/w) level in the indolinone intermediate, with a trigger for compliance action set even lower at 0.01 ppm for the sum of nitrosamine impurities. The amidation process—in which the intermediate is activated with CDI under strictly anhydrous conditions and no source of nitrite is introduced—is engineered as a confirmatory barrier, and the sunitinib free base batch is additionally sampled for liquid-liquid extraction with subsequent analysis on a triple-quadrupole mass spectrometer operated in multiple reaction monitoring mode. Packaging of the intermediate in foil-laminated, antistatic polyethylene liners inside fiber drums under a nitrogen headspace ensures that no adventitious nitrosation occurs during transcontinental shipment. The finished product in this regulatory context is a FDA-approved generic sunitinib malate capsule bearing an “AB” therapeutic equivalence rating, distributed in high-density polyethylene bottles with child-resistant closures, and the market authorization holder’s quality agreement with the intermediate manufacturer explicitly mandates an annual nitrosamine re-evaluation report per FDA CDER MAPP 5220.1.

Quality Attribute	Applicable Regulatory Standard / Guideline	Quantitative Threshold or Criterion	Routine Control Methodology
Organic impurities in starting material indolinone intermediate	ICH Q11 (starting material justification), ICH Q3A(R2)	Single unknown impurity ≤ 0.10%, total impurities ≤ 0.50%	HPLC-UV at 268 nm, C18 stationary phase, phosphate buffer (pH 3.0)/ACN gradient
Photodegradation products in sunitinib capsules	ICH Q1B (photostability), ICH Q3B(R2)	Reporting 0.2%, identification 0.5% (for max daily dose 50 mg)	Stability-indicating HPLC, resolution factor ≥ 2.0 between Z/E isomers
Nitrosamine impurities in indolinone intermediate and final API	ICH M7(R2), FDA Nitrosamine Guidance, EMA CHMP/455985/2020	Acceptable intake ≤ 26.5 ng/day; intermediate control limit ≤ 0.03 ppm; reporting threshold 0.01 ppm	LC-MS/MS (APCI, positive ion mode), multiple reaction monitoring
Residual solvents	ICH Q3C(R8)	DMF Class 2: ≤ 880 ppm; Acetone Class 3: ≤ 5000 ppm; 2-Propanol Class 3: ≤ 5000 ppm	Headspace GC-FID with DB-624 column
qNMR certified reference standard homogeneity	ISO 17034:2016, ISO Guide 35:2017, USP 〈761〉 & 〈1761〉	Assigned purity 99.5% ± 0.4% (k=2), intra-bottle CV ≤ 0.2%	Quantitative ¹H NMR (600 MHz), internal standard method

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Certification & Compliance

5-(Z)-(5-Fluoro-2-Oxoindolin-3-Ylidene)Methyl)-2,4-Dimethyl-1H-Pyrrole-3-Carboxylic Acid is manufactured under an ISO 9001 quality system and complies with relevant regulatory requirements.
COA, SDS/MSDS, and related certificates are available upon request. For certificate requests or inquiries, contact: sales9@bouling-chem.com.

More Introduction

A shipping specification for the pyrrole‑3‑carboxylic acid of the sunitinib scaffold (CAS 356068-97-8) reflects a purity of ≥98.0 % area by HPLC, with a single maximum impurity ceiling of 0.50 %. The dry, crystalline powder appears as a yellow‑to‑orange solid (lot‑dependent hue shifts ΔE ≤ 2.5 versus reference standard), and its identity is confirmed by ¹H‑NMR at 400 MHz with the characteristic methine singlet at δ 7.75 ± 0.05 and a fluorine‑coupled doublet for the indolinone H‑4 proton. Water content by Karl Fischer titration is held below 0.5 %, while residual solvents—ethanol and N,N‑dimethylformamide—are controlled to ≤5000 ppm and ≤880 ppm respectively, compliant with ICH Q3C Option 2 limits. The material is packaged under argon in amber borosilicate vials and stored at –20 °C, as ambient fluorescent light induces (Z)→(E) isomerisation at a rate approaching 2 % per 24 h when exposed to 1000 lux of cool‑white illumination. This photolability requires any handling during analytical weighing or formulation to be conducted under subdued red light, and process‑scale synthesis vessels are blanketed with nitrogen to avoid oxidative darkening that manifests as an absorbance rise at 430 nm.

What role does the free carboxylic acid serve in kinase inhibitor synthesis?

The free acid functions as the penultimate intermediate en route to the oncology therapeutic sunitinib and its analogues. After formation of the indolin‑2‑one methylene‑pyrrole core via base‑catalysed Knoevenagel condensation of 5‑fluoro‑1,3‑dihydro‑2H‑indol‑2‑one with 2,4‑dimethyl‑1H‑pyrrole‑3‑carbaldehyde, the acid is activated with a carbodiimide—most commonly N,N′‑dicyclohexylcarbodiimide or 1‑ethyl‑3‑(3‑dimethylaminopropyl)carbodiimide—and coupled to N,N‑diethylethylenediamine. The carboxylic handle obviates the requirement for transient ester hydrolysis steps that complicate the methyl ester route. In pilot‑plant runs utilising 1000 L glass‑lined reactors, a 1.05‑fold molar excess of the acid over the amine achieves a terminal amide yield of 82–88 % after recrystallisation from ethyl acetate‑cyclohexane, provided the acid input carries less than 0.2 % of the (E)‑geometric isomer. The acid is also the direct progenitor of the corresponding acyl chloride, which is employed when sterically hindered amines or anilines are grafted onto the scaffold to explore structure‑activity relationships against VEGFR‑2 and PDGFR‑β. Because the free acid retains the full conjugation of the oxindole‑pyrrole system, its UV‑visible spectrum (λ_max 430 nm in methanol, molar absorptivity ≈ 38 000 M⁻¹ cm⁻¹) serves as an in‑line process analytical technology marker: a signal drop at 430 nm follows carbodiimide activation and is used to trigger amine addition in automated feed‑back control loops.

Isomer Stability and Light‑Mediated (Z)/(E) Interconversion

The exocyclic double bond connecting the oxindole and pyrrole rings exists exclusively as the thermodynamically favoured (Z)‑configuration in the solid state, as demonstrated by single‑crystal X‑ray diffraction data collected on a Bruker D8 Venture diffractometer (Cu Kα, 100 K). However, when the compound is dissolved in polar aprotic media—dimethyl sulfoxide, N,N‑dimethylformamide, or acetonitrile—a photostationary state develops under ambient light, yielding up to 18 % of the (E)‑isomer. The (E)‑contaminant exhibits a shifted HPLC retention time under the compendial method for Sunitinib Malate (USP 43‑NF 38), eluting at a relative retention time of approximately 0.42 versus the principal peak when a 150 × 4.6 mm L1-packing column is operated at 1.0 mL/min with a phosphate buffer‑acetonitrile gradient. Process‑scale handling therefore enforces strict light‑protection protocols: reactors are equipped with sight‑glass covers, transfer lines are wrapped in aluminium‑backed tape, and final isolated product is dried under vacuum in the absence of UV‑emitting sources. Quality‑control laboratories quantifying (E)‑isomer load use amber HPLC vials and validate that the autosampler carousel temperature does not exceed 4 °C to suppress thermal‑isomerisation concurrent with analysis. For storage longer than 90 days at –20 °C, duplicate HPLC analyses on receipt and on expiry are recommended to detect any excursion beyond the 0.50 % unspecified impurity threshold.

When the 3‑carboxylic acid replaces the methyl ester in amide coupling

In classical Sunitinib synthetic routes, the methyl ester (CAS 356068‑90‑1) was the preferred building block because it avoided potential decarboxylation under the acidic conditions of the Knoevenagel cyclization. Yet adopting the free acid offers a shorter sequence by eliminating the ester saponification step and its associated yield loss—typically 4–6 %—from emulsion formation during liquid‑liquid work‑up. The trade‑off is a slightly lower solubility in the coupling solvent; the acid dissolves to ≈ 12 mg/mL in dichloromethane at 20 °C, versus ∼ 40 mg/mL for the methyl ester. To compensate, process chemists employ a pre‑activation protocol: the acid is stirred with 1.1 eq of N,N′‑carbonyldiimidazole in dry dimethylacetamide at 0–5 °C for 30 min before adding the amine. This method suppresses symmetrical anhydride formation, which is otherwise the dominant side‑product when soluble carbodiimides are applied directly. On a manufacturing scale, the acyl imidazole intermediate is not isolated; its formation is tracked by an exotherm that must be controlled within ± 2 °C of the target to avoid runaway imidazole‑catalysed polymerization of the dimethylacetamide. The resultant sunitinib free‑base crude shows a marked reduction in the RRT 0.65 by‑product that arises from residual dicyclohexylurea when the ester route is used, thereby lifting the downstream recrystallisation yield to ≥ 90 %. Direct utilisation of the acid also streamlines the preparation of active‑pharmaceutical‑ingredient (API) reference standards. The United States Pharmacopeia lists this entity as Sunitinib Related Compound B, with an acceptance criterion of ≤ 0.15 % in the drug substance monograph. Laboratories acquire milligram quantities of the pre‑certified acid to spike into system‑suitability solutions, confirming resolution between sunitinib (relative retention 1.0) and the acid (RRT 0.35 under the same gradient). The ready availability of the acid as a certified impurity eliminates the need to perform a multi‑step synthesis of the methyl ester and its subsequent hydrolysis, reducing lead time for analytical method transfer by 3–4 weeks per site.

**Table 1.** Comparative profile of key sunitinib intermediates
Property	3‑Carboxylic acid (Related Compound B)	Methyl ester	Sunitinib free base
CAS Registry Number	356068‑97‑8	356068‑90‑1	341031‑54‑7
Molecular weight (g·mol⁻¹)	314.31	328.34	398.49
Appearance (bulk)	Yellow to deep‑orange powder	Pale‑yellow microcrystalline solid	Yellow to orange crystalline powder
HPLC relative retention time^a	~0.35	~0.85	1.00
Solubility in CH₂Cl₂ at 20 °C (mg/mL)	12	40	8
Key process function	Amide‑coupling precursor; impurity standard	Protected intermediate; ICH impurity marker	Final API
Recommended storage	Amber vial, –20 °C, under argon	Amber vial, 2–8 °C	Desiccated, 2–8 °C
^a USP Monograph chromatographic conditions: L1 column, phosphate buffer pH 6.8‑acetonitrile gradient; retention times normalised to sunitinib peak.

The presence of the free carboxyl group fundamentally alters the pK_a‑dependent distribution coefficient relative to ester congeners. While published experimental dissociation constants remain sparse for this particular pyrrole‑substituted acid, computational estimation using the MarvinSketch algorithm gives a pK_a of 4.5 ± 0.3 for the ‒COOH proton, consistent with withdrawal by the flanking pyrrole ring. This acidity permits selective extraction from organic‑process streams into 0.1 M aqueous sodium bicarbonate; residual neutral esters or oxindole‑dimer impurities stay in the organic layer, providing a plant‑level polish that can reduce total organic impurities by 0.12–0.18 area%. However, exposure to aqueous base above pH 9.5 must be limited to ≤10 min at 0–5 °C, as the oxindole ring undergoes hydroxide‑catalysed ring‑opening to a 2‑aminophenylglyoxylic acid derivative (confirmed by formation of a m/z 342 peak in LC‑MS), resulting in irreversible potency loss for subsequent kinase‑inhibitor synthesis. Storage incompatibility with primary and secondary amines is critical: the free acid forms intermolecular salts with diethylamine or piperidine that crystallise as low‑solubility adducts, complicating quantitative transfer from weighing vessels. For this reason, all ancillary reagents used in conjunction with the acid are sourced as free‑base solutions in anhydrous solvents, not as hydrochloride salts, and dedicated glassware is rinsed with 0.01 M HCl followed by three changes of anhydrous acetone to remove amine residues.

**Table 2.** Regulatory and standards matrix referenced for the carboxylic acid intermediate
Standard / Guidance	Relevant section or clause	Application to the compound
ICH Q3A (R2)	Reporting, identification, qualification thresholds for impurities	Defines 0.10 % reporting threshold for unspecified impurities in batch‑release testing
USP 43‑NF 38 Sunitinib Malate Monograph	Related compounds test	Establishes the acid as Related Compound B with an acceptance criterion of ≤ 0.15 % in the API
USP General Chapter <621> Chromatography	System suitability and gradient methods	Used to set resolution requirement of ≥ 2.0 between the acid and sunitinib in the spiked standard
ICH Q3C (R8)	Residual solvents (Class 2 and Class 3)	Limits for DMF (880 ppm) and ethanol (5000 ppm) derived from synthetic route
FDA 21 CFR 211.194	Laboratory records	Mandates retention of complete analytical raw data for each lot used as an impurity standard
ICH Q1A (R2)	Photostability testing	Requires confirmatory photostability data (Option 2) under ICH Q1B for light‑sensitive intermediates

During the final amide‑bond‑forming step, process analytical control relies on a reversed‑phase UPLC method (Waters Acquity BEH C18, 1.7 µm, 50 × 2.1 mm) with a run time of 4.5 min. The acid peak at λ = 430 nm is monitored every 2 min during the activation phase; its area percent drops below 2 % of the initial value within 15 min of amine addition, signalling completion. Operators initiate the aqueous work‑up only when unreacted acid is ≤0.5 area%, as residual acid in the final sunitinib free base increases the difficulty of meeting the ≤ 800 ppm residual palladium specification when a catalytic hydrogenation path is used earlier in the sequence. Any deviation that extends the activation time beyond 40 min is investigated for possible moisture ingress in the dimethylacetamide—Karl Fischer titration of the solvent must show < 200 ppm water—because water hydrolyses the acyl imidazole and regenerates the acid, causing a deceptive plateau in the UPLC reaction‑monitoring trace.