| HS Code | 852121 |
| Productname | BOC-D-phenylalanine |
| Casnumber | 33125-05-2 |
| Molecularformula | C14H19NO4 |
| Molecularweight | 265.31 g/mol |
| Appearance | White to off-white crystalline powder |
| Meltingpoint | 85-88 °C |
| Opticalrotation | [α]D20 = -25° (c=1 in ethanol) |
| Solubility | Soluble in methanol, ethanol, DMF, DMSO and ethyl acetate; insoluble in water |
| Storageconditions | Store at 2-8 °C, protected from light and moisture |
| Purity | ≥98% (HPLC) |
As an accredited BOC-D-phenylalanine factory, we enforce strict quality protocols—every batch undergoes rigorous testing to ensure consistent efficacy and safety standards.
| Packing | BOC-D-phenylalanine is supplied as a white powder in a sealed glass bottle; net weight 25 g per container. |
| Container Loading (20′ FCL) | 20′ FCL: BOC-D-phenylalanine in sealed drums, palletized and secured, protected from moisture, with proper labeling and ventilation. |
| Shipping | BOC-D-phenylalanine ships in sealed, inert packaging under ambient conditions, protected from moisture and direct sunlight. No special hazmat designation is required for standard courier transport. Keep container tightly closed during transit and storage. Ensure compliance with local regulations for laboratory chemicals. Delivery typically takes 2–5 business days depending on destination. |
| Storage | Store BOC-D-phenylalanine in a tightly sealed container away from light and moisture. Recommended storage is refrigerated at 2–8°C. Keep in a cool, dry, well-ventilated area. Avoid exposure to strong oxidizing agents. Use proper personal protective equipment when handling. Ensure container is clearly labeled with the chemical name and relevant hazard information. |
| Shelf Life | Store tightly sealed in a cool, dry place away from light. Shelf life: typically 2–3 years. |
BOC-D-Phe is used as the N-terminal protected D-phenylalanine building block in solution-phase fragment condensation routes to somatostatin analogue peptide APIs, particularly octreotide acetate and vapreotide acetate, where the D configuration at position one confers resistance to aminopeptidase N degradation. The coupling is carried out in jacketed glass-lined reactors with anchor agitation and jacket temperature held at 0–5 °C; the carboxyl group of BOC-D-Phe is pre-activated with N,N′-diisopropylcarbodiimide and 1-hydroxybenzotriazole in anhydrous dimethylformamide for 15 min before addition to the free N-terminal amine of a protected heptapeptide fragment. The BOC-D-Phe charge is maintained at 1.05–1.15 molar equivalents relative to free amine; above 1.15 equivalents, unreacted BOC-D-Phe acid persists in the organic phase and increases the phenylalanine-related impurity signal in the crude peptide when assayed by HPLC using USP 621 and European Pharmacopoeia 2.2.46. Residual water in dimethylformamide is controlled below 0.1% because water accelerates oxazolone formation, which can epimerize the D-Phe α-carbon and produce the undesired L-Phe diastereomer. Production-scale failure modes observed during pilot campaigns include slow diisopropylurea precipitation at temperatures below 5 °C, requiring in-line filtration through 0.45 μm polytetrafluoroethylene membranes at nitrogen pressure of 0.1–0.2 MPa. After coupling, the Boc group is removed with trifluoroacetic acid, and the linear peptide is subjected to iodine-mediated disulfide bond formation in dilute acetic acid-water at approximately pH 8.0; over-oxidation to sulfoxide is suppressed by controlled iodine addition and stopped with ascorbic acid. The crude cyclic peptide is purified on preparative reversed-phase HPLC columns packed with 10 μm C18 silica, followed by acetate salt exchange and lyophilization. Terminal finished product types include octreotide acetate injection at 100 μg/mL and long-acting release microsphere intermediates for somatostatin receptor-positive neuroendocrine tumour therapy. Compliance for this downstream application is governed by ICH Q7 for active pharmaceutical ingredient GMP, ICH Q11 for starting material justification, EU GMP Part II for active substance manufacture, and residual solvent limits under ICH Q3C(R8).
In the manufacture of the peptide precursor for 177Lu-DOTA-TATE and 68Ga-DOTA-TATE radiopharmaceuticals, BOC-D-Phe functions as the N-terminal D-Phe building block in Boc/benzyl solid-phase peptide synthesis on PAM resin. The final amino acid coupling of BOC-D-Phe is performed at 4.0 molar equivalents relative to resin-bound free amine, using 2-(1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate and N,N-diisopropylethylamine in N-methyl-2-pyrrolidone for 45 min at 20–25 °C. A Kaiser test result below 0.1 absorbance units at 570 nm is used as the release threshold before the subsequent TFA deblocking step. The protected peptide chain is assembled on an automated peptide synthesizer with a 100 mmol reaction vessel; side-chain protection typically includes Lys(2-Cl-Z), Thr(Bzl), and Cys(4-MeBzl), which remain intact during repetitive TFA-mediated Boc removal. After final Boc deprotection with 50% TFA in dichloromethane, the peptide is cleaved from PAM resin with anhydrous hydrogen fluoride in the presence of anisole at 0 °C for 60 min, precipitated in cold diethyl ether, and cyclized by iodine oxidation to form the intramolecular disulfide bridge. The free N-terminal amine is then acylated with DOTA-tris(tBu) ester at 1.5–2.0 molar equivalents; chelator tert-butyl groups are removed with TFA, and the crude peptide-DOTA conjugate is purified by preparative HPLC. The terminal product is the GMP peptide precursor that is subsequently radiolabelled with 177Lu or 68Ga and formulated as an injection for neuroendocrine tumour imaging or targeted radionuclide therapy. This downstream application operates under 21 CFR Part 212 where positron-emitting radiolabelled drug production applies, EU GMP Annex 3 for radiopharmaceutical manufacturing, and ICH Q3C(R8) for residual solvent control. Analytical release of the precursor includes USP 621 for chromatographic purity, European Pharmacopoeia 2.2.46 for separation technique conformity, and bacterial endotoxin testing according to Ph. Eur. 2.6.14.
For diagnostic coagulation substrate manufacturing, BOC-D-Phe is the N-terminal building block in solution-phase synthesis of H-D-Phe-L-Pip-L-Arg-p-nitroanilide dihydrochloride, a chromogenic thrombin substrate used in clinical haemostasis testing. The coupling step charges BOC-D-Phe at 1.1 molar equivalents relative to H-L-Pip-L-Arg-p-nitroanilide, with activation by 2-(7-aza-1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate and N,N-diisopropylethylamine in dimethylformamide at -5 to 0 °C. Charge ratios below 1.05 equivalents cause incomplete acylation of the secondary amine of L-pipecolic acid, while ratios above 1.15 equivalents generate a fine suspension of residual BOC-D-Phe acid that fouls the 0.2 μm in-line filter and interferes with subsequent crystallization. The process is conducted in a Hastelloy C-22 vessel with temperature control to avoid cleavage of the nitroanilide redox label under acidic workup. After coupling, the Boc group is removed with 4 M HCl in dioxane at 20 °C, the peptide is precipitated in ethyl acetate, and the dihydrochloride salt is crystallized from ethanol-water to a residual water content below 5.0%. Terminal finished products are lyophilized chromogenic substrate reagents used in coagulation analyzers for thrombin and factor Xa activity measurements; the substrate is formulated in assay buffer at 1.0–2.0 mmol/L final concentration depending on analyzer optics. Compliance for this diagnostic application is anchored to ISO 13485:2016 for medical device quality management, IVDR 2017/746 for in vitro diagnostic reagent supply, ISO 23640:2015 for reagent stability, and USP 621 for chromatographic purity testing.
BOC-D-Phe is specified by custom peptide synthesis CRO/CDMO units when a client peptide requires an N-terminal D-Phe residue in a peptide acid and the synthetic route is based on fragment condensation rather than resin-bound Fmoc assembly. A typical 100 mmol project charges BOC-D-Phe at 1.2 molar equivalents relative to the free amine of a protected peptide fragment, with activation by N,N′-diisopropylcarbodiimide and ethyl 2-cyano-2-(hydroxyimino)acetate in dichloromethane-dimethylformamide at 0 °C for 10 min. Process equipment includes a jacketed glass reactor with a polytetrafluoroethylene-coated thermowell; in-process control samples are withdrawn at 15 min intervals for anhydrous liquid chromatography-mass spectrometry to confirm residual free amine below 0.1% before the next acylation cycle. After coupling, the Boc group is removed with trifluoroacetic acid, the crude linear peptide is precipitated in cold methyl tert-butyl ether, and the product is purified by preparative HPLC on a C18 column. Terminal finished product is a lyophilized research-grade peptide acid supplied in amber vials under argon; end users apply these peptides in receptor binding assays, cell-based assays, and stability screening. Compliance is ISO 9001:2015 for non-GMP custom synthesis, ICH Q3C(R8) for residual solvent limits, and USP 621 for final HPLC purity reporting.
When D-Phe substitution of L-Phe at the P1 position is screened for resistance to aminopeptidase digestion in early drug discovery, BOC-D-Phe is used in parallel solution-phase coupling at a fixed 1.2 molar equivalents relative to each protected peptide fragment. Couplings are executed in a nitrogen-purged 96-well reaction block with 2-(7-aza-1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate and N,N-diisopropylethylamine in N-methyl-2-pyrrolidone at 15 °C for 30 min; the crude products are precipitated with cold diethyl ether and analysed by ultra-performance liquid chromatography-mass spectrometry without intermediate purification. Published kinetic data for this specific parallel synthesis configuration is limited; the coupling stoichiometry is derived from standard solution-phase peptide synthesis protocols. Terminal finished product type is a freeze-dried preclinical peptide candidate panel used in plasma stability assays, receptor occupancy assays, and subsequent off-target profiling. Compliance is governed by ICH M3(R2) for nonclinical safety study design and ISO/IEC 17025:2017 for bioanalytical data generated to support candidate ranking.
Manufacturers of peptide impurity reference standards use BOC-D-Phe to synthesize D-Phe-containing process impurities for chromatographic system suitability testing during therapeutic peptide API release. The synthesis charges BOC-D-Phe at 1.5 molar equivalents relative to the free amine of the impurity peptide backbone, with activation by benzotriazol-1-yl-oxytripyrrolidinophosphonium hexafluorophosphate and N,N-diisopropylethylamine in dimethylformamide at 0–5 °C for 20 min. After coupling, the Boc group is removed with trifluoroacetic acid, and the crude peptide is purified by preparative HPLC, lyophilized, characterized by high-resolution mass spectrometry, two-dimensional NMR, and quantitative NMR, then dispensed under argon in glass vials as a certified reference material. Terminal finished product is a pharmacopoeial reference standard or impurity standard used in method validation and batch release of D-Phe-containing peptide APIs. Compliance for this downstream activity is ISO 17034:2016 for reference material producers, ISO/IEC 17025:2017 for characterization, and European Pharmacopoeia 2.2.46 for chromatographic purity assignment.
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BOC-D-phenylalanine, commonly catalogued as Boc-D-Phe-OH or N-(tert-butoxycarbonyl)-D-phenylalanine, is an N-protected chiral amino acid used as a building block in peptide synthesis and medicinal chemistry. The IUPAC name is (2R)-2-[(tert-butoxycarbonyl)amino]-3-phenylpropanoic acid. The substance carries CAS registry number 18942-49-9, empirical formula C14H19NO4, and molecular weight 265.31 g/mol. Commercial release specifications for research-grade material typically list chemical assay ≥98.5% by HPLC, enantiomeric purity ≥99.0%, specific rotation [α]20D −25.0° ± 1.0° (c = 1, methanol), loss on drying ≤0.5%, and residue on ignition ≤0.10%. The product is supplied as a white to off-white crystalline powder. The tert-butoxycarbonyl group protects the α-amino function under basic and nucleophilic conditions, while the free carboxylic acid permits direct activation or resin attachment. This protecting-group strategy distinguishes BOC-D-phenylalanine from Fmoc-D-phenylalanine, which is removed by secondary amines, and from Cbz-D-phenylalanine, which requires hydrogenolysis or strong acid for deprotection.
| Parameter | Method | Acceptance limit |
|---|---|---|
| Appearance | Visual | White to off-white crystalline powder |
| Chemical assay | USP <621> HPLC | ≥98.5% |
| Enantiomeric purity | Chiral HPLC | ≥99.0% |
| Specific rotation | USP <781> | −25.0° ± 1.0° (c = 1, methanol) |
| Loss on drying | USP <731> | ≤0.5% |
| Residue on ignition | USP <281> | ≤0.10% |
| Heavy metals | USP <231> | ≤10 ppm |
| Residual solvents | USP <467> | Ph. Eur. limits |
BOC-D-phenylalanine contains the unnatural D-α-carbon stereochemistry, which reverses the spatial orientation of the benzyl side chain relative to the carbamate-protected amine and the carboxylic acid. Peptide sequences that replace L-phenylalanine with D-phenylalanine are not recognized by α-chymotrypsin, trypsin, or leucine aminopeptidase, because these enzymes show stereospecificity for L-amino acid residues. In receptor-ligand design, substitution at the phenylalanine position modifies backbone trajectory and side-chain presentation without altering the functional groups. Typical enantiomeric acceptance criteria for BOC-D-phenylalanine are ≤0.1% L-enantiomer by chiral HPLC. A cellulose tris(3,5-dimethylphenylcarbamate) stationary phase with a hexane/2-propanol mobile phase and UV detection at 254 nm is sufficient to separate the enantiomers under isocratic conditions. If optical rotation is used as a screening method, USP <781> or Ph. Eur. 2.2.7 requires a path length of 1.0 dm and a thermostatted tube at 20 °C; the D-enantiomer gives a negative rotation in methanol, while the L-enantiomer gives a positive rotation of similar magnitude.
In BOC/benzyl solid-phase peptide synthesis, BOC-D-phenylalanine is coupled to aminomethyl polystyrene-divinylbenzene resin with a typical substitution of 0.5–1.0 mmol/g. Carboxyl activation is performed with 1.0 equivalent of HOBt and 1.0 equivalent of DIC in DMF or DCM at 0–25 °C for 5–10 min, followed by addition to the resin. Coupling is maintained under nitrogen with gentle agitation in a manual peptide synthesis vessel or automated synthesizer for 30–90 min. The Kaiser test is used to monitor free amine; a negative test indicates that acylation has consumed the available amino groups. If the test remains positive after a second coupling, recapping with acetic anhydride/pyridine is introduced to cap unreacted sites. BOC removal is carried out with 20–50% trifluoroacetic acid in DCM for 2 × 5 min at room temperature, and the resin is neutralized with 5–10% N,N-diisopropylethylamine in DCM. In manual synthesis at 0.25 mmol scale, incomplete neutralization can lower the next coupling efficiency by 10–20%; published data for this specific configuration is limited, but the failure mode is general to BOC SPPS.
Removal of the tert-butoxycarbonyl group proceeds through protonation of the carbamate oxygen and elimination of isobutylene and carbon dioxide. Standard TFA-based deprotection is conducted at 0–25 °C with 20–50% TFA in dichloromethane or neat TFA for final cleavage. Because the tert-butyl cation generated during BOC removal can alkylate electron-rich residues, scavengers such as triisopropylsilane, anisole, water, or 1,2-ethanedithiol are added at 2.5–5.0% v/v. BOC-D-phenylalanine itself has no electron-rich side-chain group, but scavenger use is still required when the adjacent residue is methionine, tryptophan, tyrosine, or cysteine. In BOC/benzyl SPPS, final resin cleavage with hydrogen fluoride is performed at 0 °C with anisole 10% v/v to suppress benzyl cation side reactions. The D-phenylalanine residue is stable under these conditions, but the free peptide must be precipitated from cold diethyl ether and lyophilized to avoid prolonged acid contact.
BOC-D-phenylalanine is converted to reactive intermediates for solution-phase peptide bond formation. The N-hydroxysuccinimide ester is prepared with 1.1 equivalents of N-hydroxysuccinimide and 1.1 equivalents of dicyclohexylcarbodiimide in dry DCM or THF at 0–4 °C, then allowed to warm to 20–25 °C over 4 h. Filtration removes dicyclohexylurea; the filtrate is used directly for coupling to an amino acid ester. The pentafluorophenyl ester offers similar activated stability and can be isolated. For large-scale batch reactions, the mixed anhydride method with isobutyl chloroformate and N-methylmorpholine at −15 °C is preferred because the by-products are more easily removed. Coupling with 1.0 equivalent of an amino ester in DMF at 20–25 °C generally reaches completion within 2–6 h; conversion is monitored by TLC or HPLC at 210 nm. The free carboxyl group remains available for regeneration by saponification with 1.0 M lithium hydroxide in aqueous tetrahydrofuran. Racemization is not normally observed when the carboxyl is activated as the pentafluorophenyl ester in the presence of 1.0 equivalent of N-methylmorpholine; specific published data for BOC-D-phenylalanine under all activation modes is limited.
BOC-D-phenylalanine is selected in convergent peptide syntheses when the N-terminal amine must remain protected under the basic conditions used for Fmoc removal. Fmoc-D-phenylalanine is removed with 20% piperidine in DMF at room temperature; these conditions can hydrolyze sensitive esters and promote epimerization in activated fragments. The BOC group remains intact under piperidine treatment, allowing selective removal of Fmoc at another position. Conversely, the BOC group is removed with TFA, which leaves Fmoc and Cbz intact. This orthogonality permits three-dimensional protection schemes using BOC, Fmoc, and allyloxycarbonyl groups. BOC-D-phenylalanine also has different solubility and extraction behaviour than Cbz-D-phenylalanine; the tert-butyl carbamate derivative is more soluble in ethyl acetate and dichloromethane, while the Cbz derivative is often purified by crystallization from ethyl acetate/hexane. When BOC-D-phenylalanine is used in solution-phase fragment condensations, the acid-labile BOC group must be removed before final hydrogenation if a benzyl ester is present, because catalytic hydrogenation will not remove BOC but will cleave the benzyl ester.
Chiral purity is the critical quality attribute for BOC-D-phenylalanine because the L-enantiomer produces peptide diastereomers with altered binding conformations. Routine lot release combines achiral HPLC for chemical purity and chiral HPLC for enantiomeric ratio. Achiral methods use a C18 column with acetonitrile/water mobile phases containing 0.1% trifluoroacetic acid or 0.1% formic acid, with UV detection at 210–220 nm. Chiral separation is performed on a cellulose tris(3,5-dimethylphenylcarbamate) column, with hexane/2-propanol mobile phases containing 0.1% trifluoroacetic acid. Retention times depend on column dimensions; typical differences of 1.5–3.5 min are observed between enantiomers at 0.8 mL/min on a 250 mm × 4.6 mm column. Acceptance thresholds of ≤0.5% L-enantiomer are common for exploratory research, while ≤0.1% L-enantiomer is specified for GLP or GMP peptide synthesis under ICH Q7. Optical rotation is measured according to USP <781> or Ph. Eur. 2.2.7; the specific rotation is −25.0° ± 1.0° at 20 °C in methanol at c = 1. A positive value indicates contamination with BOC-L-phenylalanine or incomplete resolution of the starting D-phenylalanine.
| Property | BOC-D-phenylalanine | Fmoc-D-phenylalanine | Cbz-D-phenylalanine | BOC-L-phenylalanine |
|---|---|---|---|---|
| Protecting group | tert-butoxycarbonyl | 9-fluorenylmethoxycarbonyl | benzyloxycarbonyl | tert-butoxycarbonyl |
| Deprotection | 20–50% TFA in DCM | 20% piperidine in DMF | Hydrogenolysis or HBr/AcOH | 20–50% TFA in DCM |
| Orthogonality | Stable to piperidine, hydrogenolysis | Stable to TFA, hydrogenolysis | Stable to TFA, piperidine | Same as D |
| Specific rotation | −25.0° ± 1.0° | Typically negative | Typically negative | +25.0° ± 1.0° |
| Main use | BOC/benzyl SPPS, solution synthesis | Fmoc/tBu SPPS | Hydrogenolytic routes | L-peptide synthesis |
| Stability under base | Stable | Labile | Labile to strong base | Stable |
| Storage | 2–8 °C | 2–8 °C | 2–8 °C | 2–8 °C |
BOC-D-phenylalanine is stable for 24 months when stored at 2–8 °C in a tightly closed container under dry nitrogen. Exposure to relative humidity above 60% increases water absorption and can reduce chemical assay over successive sampling. The product is incompatible with strong acids, strong bases, and oxidizing agents. Aqueous or alcoholic solutions should be used within 24 h if stored at 2–8 °C; prolonged standing in solution at pH outside 3–9 leads to slow carbamate hydrolysis. The free acid may be converted to the dicyclohexylammonium salt or sodium salt for improved crystallinity or solubility in polar media, but this changes molecular weight and requires stoichiometric correction. Residual solvent testing follows USP <467>, and heavy metal limits are commonly set at ≤10 ppm using Ph. Eur. 2.4.8 or USP <231> where applicable. The product is a laboratory reagent and is not intended for direct pharmaceutical administration without further purification.
At production scale, BOC-D-phenylalanine is often converted to its methyl ester hydrochloride or BOC-D-phenylalanine N-carboxyanhydride for polymerization or fragment condensation. The esterification is carried out with thionyl chloride in methanol at 0–5 °C, followed by removal of volatiles under reduced pressure at 40 °C. The N-carboxyanhydride route requires phosgene or triphosgene and is not recommended outside dedicated containment because of the acute toxicity of phosgene. BOC-D-phenylalanine is used as a starting material for D-phenylalaninol and D-phenylalanine methyl ester, which are intermediates in chiral auxiliary and Ugi multi-component reaction procedures. The product also appears in peptide-based enzyme substrates that require resistance to aminopeptidase cleavage; for example, replacing N-terminal L-phenylalanine with D-phenylalanine extends half-life in serum-containing assays, but published data for specific substrate sequences is limited. In GMP peptide production under ICH Q7, supplier qualification includes identity by infrared absorption and specific rotation, assay by HPLC, and residual solvent analysis according to USP <467>.
