COA Considerations for TB-500 | Pure Lab Peptides
Certificate of Analysis (COA) considerations for TB-500 involve verifying key quality attributes of this synthetic research peptide. TB-500 is the N-terminal acetylated 17–23 fragment of human thymosin beta-4 (sequence Ac-LKKTETQ)【19†L249-L258】. For laboratory research use, a TB-500 COA should detail the peptide’s identity (sequence and molecular weight), purity, and other analytical data. This ensures each TB-500 batch matches its specified composition under research-use-only conditions.
Fast Answer
Products discussed in this article are intended for laboratory research use only and are not intended for human or animal consumption. A TB-500 COA will report the peptide sequence (Ac-LKKTETQ), molecular weight (~888.49 Da), HPLC purity, and related data. Batch-specific COAs include test results (mass spectrometry and HPLC) confirming the peptide’s identity and purity【19†L249-L258】【2†L1638-L1643】.
TB-500 Peptide Overview
TB-500 is a synthetic peptide corresponding to residues 17–23 of human thymosin beta-4, with an N-terminal acetylation (Ac-LKKTETQ)【19†L249-L258】. In research contexts, TB-500 is supplied as a lyophilized peptide for laboratory studies. It is not intended for therapeutic use. The peptide’s COA should clearly state its sequence and theoretical molecular weight, allowing researchers to verify they have the correct compound.【19†L249-L258】
Identifying TB-500 by Mass Spectrometry
Identity confirmation relies on high-resolution mass spectrometry (MS). The expected molecular ion for TB-500 (Ac-Tβ4(17–23)) is m/z 889.4988 ([M+H]+), corresponding to a neutral mass of 888.4910 Da【19†L249-L258】. MS/MS fragmentation yields b- and y-type ions that match the known peptide sequence. According to one study, the observed fragment ions unambiguously identified the heptapeptide Ac-LKKTETQ【19†L249-L258】. The COA should list the measured mass and confirm sequence matching, often via LC–MS/MS data or a high-resolution MS spectrum.
Purity and Impurity Analysis for TB-500
Reversed-phase HPLC is the standard method for assessing peptide purity【27†L119-L128】. A COA will report the percentage area of the main peptide peak in an HPLC chromatogram. In analytical studies of TB-500, HPLC–HRMS revealed a single dominant peak with no coeluting impurities【19†L263-L270】, indicating high purity. The chromatogram itself is often included in the COA to visualize purity. Any minor peaks (related peptides or by-products from synthesis) above reporting thresholds should be noted. As per quality guidelines, specified and unspecified impurities above the identification threshold are listed on the COA with acceptance limits【2†L1638-L1643】.
| COA Component | Description |
| Peptide name/sequence | Confirms exact amino acid sequence (Ac-LKKTETQ for TB-500)【19†L249-L258】. |
| Molecular weight (MS) | Measured by MS; expected ~888.49 Da for Ac-Tβ4(17–23)【19†L249-L258】. |
| HPLC Purity | Percentage of main peptide peak in HPLC (typically ≥95%). A clean chromatogram peak indicates few impurities【19†L263-L270】【27†L119-L128】. |
| Amino acid analysis | Post-hydrolysis amino acid quantitation to verify peptide composition【32†L59-L66】【42†L7-L12】. |
| Other (Water/Counter-ions) | Moisture content (Karl Fischer) and any counter-ion (e.g. acetate) are measured as per guidelines【42†L19-L23】. |
COA Content and Quality Criteria
The Certificate of Analysis for TB-500 should include all critical test results. This typically covers the peptide’s sequence and name, molecular formula, measured mass, purity percentage, and a copy of the analytical chromatogram. It should specify the analytical methods used (e.g. RP-HPLC conditions, MS settings) and compare results to acceptance criteria. Official guidance suggests amino acid analysis and/or complementary methods to confirm sequence【42†L25-L28】【42†L7-L12】. Any counter-ions or salts (such as acetic acid) present are also reported【42†L19-L23】. Clear batch documentation allows researchers to cross-check each lot of TB-500 against specification limits, enhancing experimental reliability.
Research-Use Quality and Documentation
For RUO compliance, all COA data must be interpreted strictly in a laboratory research context. Researchers should verify that TB-500 batches meet stated criteria before use. Analytical reports (HPLC, MS, AAA) should align with the values on the COA. As EMA guidance notes, using multiple tests (MS, NMR, or AAA) is important to unambiguously confirm peptide structure【42†L25-L28】. By comparing batch-specific COAs, researchers ensure consistency and traceability. In practice, a comprehensive COA helps prevent use of misidentified or impure material in preclinical studies.
FAQs
What is TB-500?
TB-500 is a research peptide corresponding to a fragment of human thymosin beta-4 (Ac-LKKTETQ, amino acids 17–23)【19†L249-L258】. It is supplied for laboratory research only. The COA for TB-500 confirms the peptide’s sequence and purity, ensuring researchers know they have the correct peptide according to published specifications【19†L249-L258】.
Why is a COA important for TB-500 research?
A COA provides verified analytical data for each TB-500 batch. It confirms identity (via mass spectrometry) and purity (via HPLC) against defined specifications【19†L249-L258】【2†L1638-L1643】. For researchers, the COA ensures the peptide sample matches its claims, which is critical for reproducible experiments. Without a COA, the composition of TB-500 cannot be independently confirmed.
How is TB-500 identity confirmed?
Identity is confirmed by mass spectrometry. Analysts verify that the measured molecular mass matches ~888.49 Da (Ac-Tβ4(17–23)) and that MS/MS fragments align with the expected sequence【19†L249-L258】. The COA typically lists the observed m/z values and confirms they match the theoretical values for TB-500, ensuring the peptide’s identity.
How is TB-500 purity assessed?
Purity is assessed by reversed-phase HPLC. A high-performance liquid chromatography run should show a single main peak for TB-500, with minimal side-peaks. The area under the main peak (usually >95% in research-grade material) is reported as purity【19†L263-L270】【27†L119-L128】. The HPLC chromatogram itself is often included on the COA as evidence of this purity.
Is amino acid analysis used for TB-500?
Yes. Amino acid analysis (AAA) can be used to confirm TB-500’s composition by quantifying its constituent amino acids【32†L59-L66】. EMA guidelines note that AAA usually complements peptide characterization【42†L7-L12】. In practice, AAA provides an additional check on peptide content or sequence identity alongside MS and HPLC data.
Next Steps
Review batch-specific documentation before selecting any research-use-only peptide. Pure Lab Peptides provides detailed COAs and clear labeling for TB-500, enabling researchers to confirm identity and purity. For reliable peptide sourcing, choose suppliers who offer transparent labelling and comprehensive analytical data.
References
- Esposito S, Deventer K, Goeman J, Van der Eycken J, Van Eenoo P. “Synthesis and characterization of the N-terminal acetylated 17-23 fragment of thymosin beta 4 identified in TB-500, a product suspected to possess doping potential.” Drug Test. Anal. 2012;4(9):733-738. doi.org/10.1002/dta.1402
- Vergote V, Burvenich CPG, Van de Wiele C, De Spiegeleer B. “Quality specifications for peptide drugs: A regulatory-pharmaceutical approach.” J. Pept. Sci. 2009;15(11):697-710. doi.org/10.1002/psc.1167
- Mant CT, Chen Y, Yan Z, Popa TV, Kovacs JM, Mills JB, Tripet BP, Hodges RS. “HPLC analysis and purification of peptides.” Methods Mol. Biol. 2007;372:1-33. doi.org/10.1007/978-1-59745-430-8_1
- Qasrawi DO, Petrotchenko EV, Borchers CH. “Amino acid analysis for peptide quantitation using reversed-phase liquid chromatography combined with multiple reaction monitoring mass spectrometry.” Anal. Bioanal. Chem. 2023. doi.org/10.1007/s00216-023-04840-2
- European Medicines Agency. “Guideline on the development and manufacture of synthetic peptides.” EMA Scientific Guideline. 2013. ema.europa.eu