Peptide Analytical Testing: Purity, Sterility, and Endotoxin

Last reviewed: October 2026 · Cleared Peptides research team

Research use only. Peptide Analytical Testing: Purity, Sterility, and Endotoxin is supplied for laboratory research. It is not a drug, food, cosmetic, or dietary supplement. It is not FDA-approved. It is not for human or veterinary administration. This page does not give a dose or a method of administration. A study cited on this page describes that study’s own test article. It does not make this lot an approved medicine. To check a fact, open the PubMed or DOI link, confirm the authors and year, and match the paper’s compound name, CAS number, or sequence to the specification table. The full method is on the research desk page. Identity and purity for a lot are only what that lot’s certificate of analysis reports. An HPLC percentage is not a sterility result and not an endotoxin result unless the certificate prints those tests.

A peptide certificate is easier to read when each test is kept separate. HPLC purity, identity, net peptide content, a sterility test or sterility screen, and an endotoxin test answer different questions. A high purity percentage does not establish that a vial is sterile, free of endotoxin, or equal to the labeled milligrams. This guide explains what those tests entail, the equipment laboratories use, the written standards behind them, and how to judge a laboratory report in the United States and elsewhere.

The pages in this library are educational. They do not provide dosing, administration, or therapeutic instructions. A research catalog material is not an FDA-approved drug product. The tests below describe analytical and microbiological quality control, not a medical use.

In this guide:

• What a report covers
• HPLC purity test
• Identity and mass spectrometry
• Net peptide content
• Sterility test
• Endotoxin test
• Other tests on a peptide COA
• Standards behind the tests
• Laboratories: capable reports and weak reports
• Recognition in the United States
• FAQ

What a peptide testing report covers

A certificate of analysis (COA) is the laboratory record for one sample. Match the product name, sample identifier, and lot number to the label before reading any number. Then list the methods that were actually performed. If sterility or endotoxin is not on that certificate, the report does not establish them, even when a different lot was tested for those items.

Cleared Peptides publishes lot certificates on the COA page. Coverage varies by certificate. Some reports include HPLC purity, a measured amount, and an identity method such as FTIR or mass spectrometry. Some also include an endotoxin result and a sterility screen. The worked example on the COA page is explicit that one BT Labs report of retatrutide covers identity, purity, potency, and composition, and that it does not report particulate matter, microbial contamination, or endotoxins. Read the scope lines on the certificate in front of you.

HPLC purity test

The purity test on most peptide certificates is reversed-phase high-performance liquid chromatography with ultraviolet detection. The laboratory dissolves a portion of the sample, injects it, and separates components on a hydrophobic column. The chromatogram is integrated. Purity is the main peak’s area divided by the total area of the peaks the method detects, expressed as a percent. That figure is often called area-percent purity.

Typical equipment is a liquid chromatograph: a pump that delivers a controlled gradient, a degasser, an autosampler or manual injector, a column oven, a reversed-phase column, an ultraviolet or diode-array detector, and chromatography software. Peptide bonds absorb near 214 nm, so 214 nm is the usual detection wavelength. 280 nm is useful when the sequence contains tryptophan or tyrosine, and it can miss impurities that lack those residues. Columns are commonly C18. More hydrophobic or lipidated peptides, including acylated GLP-1 analogs, sometimes need a less retentive C8 or C4 column, a higher acetonitrile fraction, or a higher column temperature so the main peak elutes as a single, integrable peak.

Mobile phases are usually water and acetonitrile with an ion-pairing acid such as trifluoroacetic acid or formic acid. The acid sharpens peaks and helps retain short, very polar peptides that would otherwise wash off a C18 column. Short Khavinson-type peptides of two to four residues are a common example. The written method should name the column, flow rate, gradient, detection wavelength, and sample concentration. Without those conditions, two “99%” results are not comparable.

Area-percent HPLC does not see everything in the vial. Water, most salts, and many counterions (acetate, trifluoroacetate) have little or no absorbance at 214 nm, so they do not lower the purity percentage even though they add mass. A purity result is also not an identity result: an impurity with the same retention time can hide under the main peak, which is why mass spectrometry is paired with chromatography. USP general chapter <621> Chromatography and European Pharmacopoeia method 2.2.29 describe how chromatographic tests are performed and system suitability is judged. They do not, by themselves, set a purity limit for a research peptide. Any acceptance criterion, such as “not less than 98%,” has to be printed on the specification or the certificate.

Identity: mass spectrometry, and what FTIR adds

Identity testing asks whether the material is the compound named on the label. For a peptide, the strongest routine answer is a mass measurement. Electrospray ionization mass spectrometry, often coupled to the chromatograph as LC-MS, or matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF), reports a molecular mass. The analyst compares it with the mass calculated from the sequence, including known modifications such as a fatty-diacid chain on semaglutide or an Aib substitution. A mass that matches is strong evidence of identity. It can still miss some isomers that share a mass, so sequence-level questions sometimes need tandem MS (MS/MS) or amino-acid analysis.

Equipment is a mass spectrometer with an electrospray or MALDI source, a mass analyzer (quadrupole, time-of-flight, or an ion trap), a vacuum system, and software that deconvolutes the charge states of a multiply charged peptide into one intact mass. LC-MS adds the chromatograph described above, usually with formic acid rather than trifluoroacetic acid because TFA can suppress electrospray signal.

Some certificates, including examples published for this catalog, use Fourier-transform infrared spectroscopy (FTIR) as the identity method. FTIR records an infrared fingerprint of the sample, including excipients if they are present. It is a real spectroscopic test and a useful consistency check. It is less specific than a mass measurement for distinguishing closely related peptide sequences. Read the wording on the report: “consistent with” an FTIR library spectrum is not the same sentence as “monoisotopic mass matches the theoretical mass.”

Net peptide content is not the purity percentage

Net peptide content is the fraction of the powder that is peptide, as opposed to water, counterion, and residual solvent. HPLC purity can be 99% while the vial’s powder is mostly salt and moisture, or the measured peptide amount can differ from the labeled amount. Amino-acid analysis and an HPLC assay against a well-characterized reference standard are the usual content methods. Karl Fischer titration measures water. Ion chromatography measures counterions such as acetate or trifluoroacetate.

On the certificates illustrated on our COA page, “content/potency” is the measured peptide amount compared with the labeled amount, and a separate mass includes excipients. Those are different numbers. Do not multiply a purity percentage by the gross fill weight and treat the result as the peptide amount unless the certificate already reports a measured peptide amount.

Sterility test, and how it differs from a PCR screen

A sterility test asks whether viable microorganisms can be recovered from the sample under a stated method. It does not measure chemical purity, and a passing result is not a license for human or veterinary use.

The compendial method is USP <71> Sterility Tests, aligned with European Pharmacopoeia 2.6.1. The sample is either filtered through a membrane that is then cultured, or it is inoculated directly into media. Two media are used so that different organisms can grow: fluid thioglycollate medium for anaerobic and some aerobic bacteria, and soybean-casein digest medium for aerobic bacteria and fungi. The compendial incubation is 14 days, with examination for turbidity. The work is done in a clean-air environment, typically a laminar-flow hood or an isolator, with growth-promotion tested media and incubators held at the temperatures the chapter specifies. A conventional microbiology laboratory doing this test has membrane-filtration manifolds, sterility-test canisters, incubators, and environmental monitoring. It is a slow test by design.

A sterility screen by PCR, which is the screening method named on Cleared Peptides’ comparison of certificate coverage, is a different test. Polymerase chain reaction, or a quantitative PCR instrument, copies microbial DNA so it can be detected with primers aimed at bacterial or fungal sequences. Equipment is a thermal cycler, reagents, and controls that show the reaction worked. PCR can be finished in hours. It detects nucleic acid, including DNA from dead cells, and it only detects organisms that the primers can amplify. A PCR screen is not a completed USP <71> sterility test. A certificate should say which of the two was done. “Sterile” without a method name is not a result.

Lyophilized research peptides are chemical reference materials unless a specific sterile manufacturing process is documented. Absence of a sterility line on a certificate means sterility was not established for that lot.

Endotoxin test

Endotoxin is lipopolysaccharide from the outer membrane of Gram-negative bacteria. It is not the same thing as a living microbe. A sample can be sterile and still contain endotoxin, and a low endotoxin result does not prove sterility. Parenteral drug products are tested for endotoxin because of that distinction. Research certificates sometimes include the test for the same analytical reason. The result is still not a clinical clearance.

The compendial method is the bacterial endotoxins test, USP <85> and European Pharmacopoeia 2.6.14. It uses a lysate of horseshoe-crab amebocytes (LAL) that clots or changes color in the presence of endotoxin. Three formats are in use. Gel-clot reports a pass or fail at a chosen sensitivity. Chromogenic and turbidimetric kinetic methods measure a change in absorbance over time on an incubating microplate reader, commonly around 405 nm for the chromogenic substrate, and convert that time to endotoxin units using a standard curve. The standard is a control standard endotoxin calibrated against a reference standard. Glassware and diluents have to be depyrogenated, because a dirty tube can create a false positive. Cartridge systems, such as those used with a handheld or benchtop LAL reader, are a packaged form of the same chemistry.

USP <86> Bacterial Endotoxins Test Using Recombinant Reagents is the animal-free counterpart. It uses recombinant Factor C, or a recombinant cascade, instead of lysate prepared from horseshoe crabs. The equipment is similar: an incubating reader, a calibrated endotoxin standard, and endotoxin-free consumables. A report should name which chapter or method was used and the unit basis, usually EU per milligram or EU per milliliter. The FDA’s 2012 guidance “Pyrogen and Endotoxins Testing: Questions and Answers” explains how these tests are used in drug manufacturing. It does not create an endotoxin limit for research-use catalog peptides. If a certificate prints a specification, that printed limit is the one to compare with the result.

Other tests that show up on peptide certificates

• Appearance. A visual check against the specification, such as “white to off-white lyophilized powder.” This uses a viewing cabinet or a simple inspection under stated light. It does not measure purity.
• Heavy metals. Inductively coupled plasma mass spectrometry (ICP-MS) reports elements such as lead, arsenic, cadmium, and mercury. The instrument is a plasma source and a mass spectrometer. Copper peptides are a special case: copper is part of the intended complex, so a copper result has to be read against the claimed stoichiometry rather than treated as a contaminant by default.
• Residual solvents. Gas chromatography, often headspace GC, looks for solvents left from synthesis. USP <467> is the compendial framework used for drug substances.
• Peptide-related impurities. Deletion sequences, incomplete deprotection, oxidation of methionine, cysteine, or tryptophan, and deamidation of asparagine or glutamine. HPLC and LC-MS are the tools. Oxidizable residues are also why storage conditions matter; see the storage and handling guide.
• Targeted chemical screens. Some lots are additionally screened for specific unrelated compounds, such as a fentanyl screen, when that line appears on the certificate. A screen is only as broad as the analyte list printed on the report.

Standards of testing

Standards say how a competent laboratory behaves. They are not a product approval.

Metrological traceability, in the NIST sense, means a measurement is linked to a reference through a documented chain of calibrations. A lot number is not that chain. When a result matters for a regulated study, ask the laboratory for the method, the calibration, and the accreditation scope.

Which laboratories are in a strong position, and which reports are weak

There is no honest universal list of “good peptide labs” and “bad peptide labs” that stays true as ownership, methods, and accreditation scopes change. What can be checked on any report is the laboratory’s independence, its method, and whether a recognized body has assessed that method.

A report in a strong position has most of the following. The laboratory is named as a legal entity, with an address. The sample identifier and lot can be matched to the vial. Each result names a method. Chromatographic purity includes the conditions or offers the chromatogram. Identity is a mass or another stated spectroscopic comparison, not a sentence that only repeats the product name. Acceptance criteria are printed next to results. The report date, a unique report number, and a way to confirm the document with the issuing laboratory (a portal, a QR code that resolves to the laboratory, or a direct reply from the laboratory) are present. If the laboratory claims ISO/IEC 17025, the accreditation body’s certificate number is on the report or the directory, and the published scope includes the technique you are relying on. HPLC on a scope does not automatically cover sterility or endotoxin.

A weak report is one you cannot check. Typical signs: no laboratory name; a purity percentage with no method and no chromatogram; the same chromatogram image reused across unrelated compounds; “pharmaceutical grade” or “FDA approved” with no approval number; a sterility claim from a package that only contains HPLC; a lot line left unspecified when you need to match a vial; and no way to reach the laboratory that supposedly signed the page. An in-house chromatogram from the seller can be useful process information. It is not the same document as an independent laboratory’s certificate. Geography is not a substitute for any of this. A laboratory in the United States is not capable because of its country, and a laboratory in Germany or the Czech Republic is not incapable because it is outside the United States.

Types of laboratories you will actually encounter:

• Independent analytical laboratories. These are the usual issuers of a third-party peptide COA. Quality turns on the items above, especially accreditation scope.
• Manufacturer quality-control laboratories. Appropriate for in-process checks. Prefer an independent report when the certificate is the public evidence for a lot you did not make.
• University and core facilities. Often well equipped for HPLC and mass spectrometry. Accreditation varies; ask for the method and whether the work was done under a quality system.
• Clinical laboratories. In the United States these are certified under CLIA to test human specimens. CLIA is the wrong framework for the chemical identity of a research vial.
• Government laboratories (for example FDA field laboratories, or a state public-health laboratory) serve regulatory and public-health programs. They are not the typical vendor of a catalog peptide COA.

Which laboratories are recognized in the United States

The FDA does not publish a roster of approved peptide-testing laboratories for research-use materials, and it does not “approve” a laboratory in the way a certificate sometimes implies. FDA registers drug establishments and inspects manufacturing when the product is a drug. An FDA establishment registration is not an ISO/IEC 17025 accreditation, and neither one is an approval of a research peptide.

The recognition that is specific to testing laboratories is accreditation to ISO/IEC 17025 by an accreditation body that is a signatory to the International Laboratory Accreditation Cooperation mutual recognition arrangement (ILAC MRA). In the United States, bodies that accredit testing laboratories under that system include:

• A2LA (American Association for Laboratory Accreditation), with a public directory of accredited organizations.
• ANAB (ANSI National Accreditation Board).
• IAS (International Accreditation Service).
• NVLAP, the National Voluntary Laboratory Accreditation Program at NIST.
• PJLA (Perry Johnson Laboratory Accreditation, Inc.).

Search the directory, open the scope, and confirm that the method on your certificate is listed. A laboratory can be accredited for environmental metals and not for peptide HPLC. Accreditation also expires; the directory date matters.

Outside the United States, the same ILAC system is used by national accreditation bodies. DAkkS is the body in Germany. The Czech Accreditation Institute (CAI) is the body in the Czech Republic. A German or Czech report can be in the same position as a US report when the accreditation number and scope check out. Cleared Peptides’ certificate sources include US, German, and Czech laboratories; the country line is not the quality line.

Two US programs that are often confused with this:

• CLIA, run through CMS, certifies laboratories that examine human specimens for diagnosis, prevention, or treatment. A CLIA certificate does not establish competence to release a peptide purity result.
• DEA registration is required for activities with controlled substances. It is not a peptide-testing accreditation.

USP writes the chapters cited above. USP does not license laboratories. Citing USP <85> on a certificate means the laboratory claims to have followed that chapter, which you can judge from the method description and, when it matters, from the accreditation scope.

A practical order for reading a lot record

1. Match the laboratory, report number, product name, and lot or sample identifier to the vial. If the lot is printed as unspecified, the certificate alone cannot tie itself to a vial.
2. List the methods. Note which of purity, identity, content, sterility, and endotoxin are present and which are absent.
3. Read HPLC purity as area percent under the stated conditions. Read any measured milligrams as a separate result.
4. If accreditation is claimed, look up the body and the scope.
5. Confirm the report with the issuing laboratory when the result will be relied on.
6. Keep the PDF with the laboratory records for that lot. Storage after receipt is covered in the handling guide.

Compound guides in this library give the sequence, formula, and mass a laboratory would compare with an identity result. Start from the research library, for example BPC-157, semaglutide, or Epitalon.

Frequently asked questions

What does a peptide purity test measure?

On a typical certificate it is HPLC area-percent purity: the main peak’s share of the ultraviolet signal the method detects. It does not measure sterility, endotoxin, or the milligrams of peptide in the vial.

What is a sterility test?

USP <71> is a 14-day culture test, by membrane filtration or direct inoculation, in fluid thioglycollate medium and soybean-casein digest medium. A PCR sterility screen detects microbial DNA on a thermal cycler and is not the same test. The certificate has to say which one was performed.

What is an endotoxin test?

It measures bacterial endotoxin activity in endotoxin units, using LAL under USP <85> or a recombinant Factor C method under USP <86>. Equipment is an incubating reader or a cartridge reader, a calibrated endotoxin standard, and endotoxin-free labware. Endotoxin is separate from sterility.

Which laboratories are approved in the United States?

The FDA does not approve research-peptide testing laboratories. Look for ISO/IEC 17025 accreditation by an ILAC MRA signatory such as A2LA, ANAB, IAS, NVLAP, or PJLA, and confirm that the certificate’s methods are inside the published scope. CLIA is for clinical testing of human specimens, not for this use.

Does a third-party COA make a research peptide safe to consume?

No. These materials are supplied for laboratory research. A certificate describes the tests that were run. It is not FDA approval and it is not a direction for human or veterinary use.

Selected references

Research Use Only Notice

This guide is provided for educational and laboratory reference purposes. It describes analytical and microbiological tests used to characterize research materials. It does not describe, recommend, or imply any use in humans or animals. Products sold by Cleared Peptides are for laboratory research use only; they are not drugs, foods, cosmetics, or dietary supplements, are not FDA-approved, and are not intended to diagnose, treat, cure, mitigate, or prevent any disease. No dosing, administration, or therapeutic information is provided.