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A General Guideline for Storage & Handling of Catalog Peptides

A General Guideline for Storage & Handling of Custom Peptides

Frequently Asked Questions about Peptides

Frequently Asked Questions about Peptide Synthesis

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A General Guideline for Storage & Handling of Catalog Peptides
Peptide Storage and Shipping

Q.    Where can I get my Analytical Data Sheet or Certificate of Analysis?
A.    Supporting analytical information is reported in the Analytical Data Sheet (ADS) for each product.

Q.    What about the Safety Data Sheet?
A.    The Safety Data Sheet (SDS) for a each product may be found on each product's page as a PDF to download.  In the cases where the SDS has not been loaded yet, or is not online, they may be obtained by request via email. Complete physical, chemical, biological, pharmacological or toxicological properties of the products are not known. The products are intended to be used by qualified professionals under proper laboratory safety practices in appropriate facilities.

Q.    What are the shipping conditions for the peptide I am receiving?
A.    Most peptides are shipped at room temperature, and peptides are delivered to you in a lyophilized state. These shipping conditions do not compromise quality since these products are stable at room temperatures and above for as long as several weeks. Once a product arrives at your facility, it should be kept at -20 °C for long-term storage.  In a rare instance where a peptide requires shipment at low temperature, a dry ice shipment will be arranged. The cost of the shipment is the responsibility of the customer.

Q. How should I store my peptide?
A.    •    Store in a desiccator or with a desiccant to maintain a dry environment.
        •    Store in the freezer at a temperature of -20 °C

Q.    What is the appearance of my peptide?
A.    The physical state for peptides ranges from amorphous solid to crystalline powder.  Some peptides will appear as a small disk at the bottom of the vial, others may appear fluffy or powdery, and other peptides may not seem visible at all.  Particularly, peptides ordered in small quantities such as 0.1 mg or 0.5 mg, may not be visible, especially through an amber glass vial.

Note: Peptides supplied as white lyophilized (freeze-dried) powder can differ in visual appearance between vialed lots due to various components in the process. For example, during exposure to nitrogen flush, the peptides can settle in different patterns. The freezing cycle can also contribute to visual differences depending on the rate; a slower rate will compact the peptide into a dense form. During the drying process, more water may evaporate, leaving the very little peptide visible to the naked eye. The pressure applied in the process will differ from lot to lot as well. These possible scenarios may change the appearance of the white mass in the vial but does not change the amount of peptide contained in the vial.

Q.    How stable is the peptide I received?
A.    In general, peptides are quite stable.  In addition, synthetic peptides, unlike proteins purified from cells, have an extremely low chance of proteinase contamination. 
Peptides are stable for more than one year if they are stored in a lyophilized state at -20 °C or below and protected from moisture and light. However, following reconstitution of a peptide in solution, stability and storage time will decrease.
If a peptide has been in solution for an extended period of time, homogeneity of the peptide must be reconfirmed. Typically, once a peptide is in solution, it should be used within a few weeks, even when stored below -20 °C. For specific information on storage time of a peptide solution, refer to the product sheet that  comes with the order.

Conditions that can affect peptide stability include the following:

  • Contamination from microorganisms or metal ions can lead to peptide-bond cleavage. Use sterile buffer or water to reconstitute the peptide.
  • Moisture can lead to hydrolysis of the peptide. The peptide should be allowed to warm gradually to room temperature in a desiccator to reduce condensation of water vapor.
  • Constant freezing and thawing can compromise peptide integrity; therefore, stock solutions should be aliquoted.
  • O2 can negatively affect Trp, Cys, and Met residues in a peptide. If a peptide contains any of these residues, Peptides International will blanket the peptide in argon prior to sealing the vial, or screw-cap bottle.
            •       Certain amino acid bonds in a peptide are more problematic:
                        Asp-Pro bonds are sensitive to acid cleavage.
                        Asn-Gly and Asp-Gly bonds, and sometimes Asp N-term to short side-chain residues (i.e. Ser, Thr, Ala, Asn), can cyclize to form an
                        aspartimide intermediate which, in turn, can undergo spontaneous changes that may alter the peptide.

Q.    If my peptide is hygroscopic in nature, how should it be handled?
A.    A hygroscopic peptide contains charged amino acids (ex: Arg, Asp, Glu, His, Lys), making it vulnerable to exposure to oxygen which can lead to moisture uptake. To prevent the product from liquefying during the weighing process, it should be allowed to warm to room temperature in a desiccator prior to weighing.
If possible, weigh in a glove box to prevent exposure to oxygen and blanket vial with an inert gas prior to restorage. If this is not an option, weigh product quickly and close lid tightly to reduce exposure to the air. If you do not have ideal weighing conditions for this product, it is best to purchase smaller vial sizes to avoid storage after opening.

Q.    How should I reconstitute a peptide in solution?
A.    To ensure peptide integrity, these recommendations should be followed:

  1. The peptide should be allowed to warm gradually to room temperature in a desiccator in order to minimize condensation of water vapor upon opening the vial or screw-cap bottle.
  2. Visually locate the peptide in the container. Tap the vial (or vortex) to release any product that may have become trapped in the cap.
  3. Follow the instructions for solubility that accompanies the product. Products that are sold as  –s or  –v have lot-specific solubility sheets that accompany each vial.
  4. Use sterile buffer or water to reconstitute the peptide. For smaller quantities such as 0.1 or 0.5 mg, it is recommended to use a sterile syringe to inject solvent into the vial as opposed to opening the cap. If DMSO is required for solubilization, be sure to use analytical grade DMSO. DMSO can degrade and become dilute with time because it can take up water from the air, so care should be taken when using stock DMSO.
  5. Aliquot the remaining peptide solution into single-use sterile glass or high quality polypropylene vials for storage to prevent repeated freezing and thawing, which can be detrimental to the integrity of the peptide.
  6. Store stock solution in a freezer at -20 °C or below under dry conditions with a pH of 5-7.

Q. What is peptide purity?
A. This is the percentage of peptide that is found in the correct sequence as opposed to truncated, deleted, or incomplete sequences that can arise from peptide synthesis. The purity is determined by high performance liquid chromatography (HPLC).

Q. What is the difference between gross peptide weight and net peptide weight?
A. Net peptide weight is the weight of the total peptide. Some products from the Peptide Institute are distributed in pre-measured vials. These are indicated by the suffix –s or –v in the catalog. The net peptide weight is precisely determined by amino acid analysis after acid hydrolysis, HPLC, and/or UV absorption, and the value is indicated clearly on the vial label. The indicated weight is only for the net peptide molecule, and the weights of any constituents are excluded from the quantity. The amount of usable peptide meets and may exceed its advertised quantity.

Gross peptide weight is the weight of the peptide and peptide impurities as well as non-peptide components, such as water and accompanying salts. Peptides manufactured by Peptides International are sold in gross peptide weight.

Q. Can you explain the packaging of the the Peptide Institute products (bulk, -v, -s products)?
A. We carry the full line of quality products of the Peptide Institute of Osaka, Japan.  Those that have a catalog number with a  -v and -s suffix are packaged as net peptide and are in injection vials.  The products without a suffix are "bulk" and packaged by gross weight.

Vialed (-v and -s) Products:

A peptide with constant weight is lyophilized and sealed under nitrogen in each vial. The net peptide weight is precisely determined by amino acid analysis after acid hydrolysis, HPLC analysis and/or UV absorption measurement, which is indicated clearly on the label of each vial. The indicated weight is only the net peptide molecule and does not include the weight of any additional constituents (water, acetic acid, and so on). For example, code 4002-v Bradykinin is described as follows:

PBK-4002-v Bradykinin (0.5 mg vial), M.W. 1060.2 g/mol

This indicates that each vial contains approximately 0.5 mg of bradykinin and the exact weight is indicated on the label (for example, 0.53 mg) and the instruction sheet (for example, 0.53 mg, 0.50 μmol). The weight is determined carefully and precisely by experts in our quality control department, therefore, we guarantee the quantity in each vial even if the content seems to be a small quantity.

A peptide solution of a known concentration can be constituted easily by injecting a given volume of a suitable solvent, indicated in the instruction sheet, into the vial using a calibrated syringe and dissolving the contents thoroughly. The peptide content in each vial is relatively small and accurately measured, therefore, the peptide should not be taken out of the vial to prepare a solution with a guaranteed concentration. The instruction sheet can be found on the last page of the ADS/COA.

Bulk Products:

The amorphous powder of each peptide is thoroughly dried over desiccant in vacuo and then weighed into a screw-capped bottle. Thus, the weight indicated on the label represents the gross weight of the amorphous powder, which includes the peptide as well as the accompanying water and acetic acid, if any. This is also called "gross" peptide. The amount of water and acetic acid in each amorphous powder is precisely determined by elemental analysis, Karl Fischer titration or gas chromatography. In some cases, peptides contain hydrochloride, trifluoroacetate, or ammonia instead of acetic acid. The observed value(s) of such accompanying constituent(s) is given in the structural formula of the respective peptide described in this catalog. For example, code 4002 Bradykinin is described as follows:

PBK-4002 Bradykinin (Bulk 100 mg)

M.W. 1060.2 • 120.10 • 54.05 g/mol

The total molecular weight of this amorphous powder is calculated to be 1234.4, which consists of 1060.2 for the net bradykinin molecule, 120.10 for two molecules of acetic acid and 54.05 for three molecules of water. This means that 100 mg of this powder contains net 85.9 mg of bradykinin molecules. Amounts of the accompanying water and acetic acid vary with the lot; the exact value in the purchased peptide is available on request. Given the hygroscopic nature of amorphous powder, precise weighing of a small quantity of peptide is not an easy task.

Liability Disclaimer
All products sold by Peptides International, Inc. are intended solely for laboratory and research use and should not be used in or on human subjects.  User assumes all risk of patent infringement by reason of use of material provided by Peptides International. Peptides International will not be responsible for damages arising from misuse of any product and is not responsible for the results of research using our products. 

 


Custom Peptide Guidelines FAQs:

A General Guideline for Storage & Handling of Custom Peptides

Q.    How stable is a peptide?
A.
    Most peptides experience very little degradation over time and are stable for more than one year if they are stored in a lyophilized state at -20 °C or below and protected from moisture and light.  However, following reconstitution of a peptide in solution, stability and storage time will decrease.  We do not test the stability of a peptide solution; therefore, prompt use is recommended.  However, if you decide to store a peptide solution for several weeks, you should aliquot out the solution into clean, inert glass or plastic vials to prevent freeze-thaw cycles.

Conditions that can affect peptide stability include the following:

•    Contamination from microorganisms or metal ions can lead to peptide-bond cleavage.  Use sterile buffer or water to reconstitute the peptide.
•    Moisture can lead to hydrolysis of the peptide.  The peptide should be allowed to warm gradually to room temperature in a desiccator to reduce
     condensation of water vapor.  
•    Constant freezing and thawing can compromise peptide integrity; therefore, stock solutions should be aliquoted
•    Peptides containing Cys or Met are susceptible to oxidation due to the side chain groups with oxygen.  It is advisable to blanket the peptide with
     argon or nitrogen when the vial is opened. Buffers used to dissolve these peptides should be degassed, either by bubbling argon or nitrogen
    through the solution for 10 minutes, or by subjecting the solution to high vacuum for 10 minutes using a common ultrafiltration capsule.  Peptides
     containing such amino acids tend to have very short-term stability, and long-term storage is not recommended.  In some cases, peptides containing
     Trp may be hygroscopic and require similar handling methods.
•    Certain amino acid bonds in a peptide are more problematic:
    ○    Asp-Pro bonds are sensitive to acid cleavage.
    ○    Asn-Gly and Asp-Gly bonds, and sometimes Asp N-term to short side residues (Ser, Thr, Ala, Asn), can cyclize to form an aspartimide
          intermediate which, in turn, can undergo spontaneous changes that can alter the peptide.

Q.    What are conditions for shipping for a custom peptide?
A.    Custom peptides are shipped at room temperature and delivered in a lyophilized state.  These shipping conditions do not compromise quality since these products are stable at room temperatures and above for as long as several weeks.  Custom peptides containing Cys or Met are susceptible to oxidation due to the side chain groups with oxygen.  These peptides are packaged under argon gas in order to displace oxygen and reduce likelihood of oxidation.  In some cases, peptides containing Trp may be hygroscopic and require packaging under argon gas as well.

Q.    Where can I get my Analytical Data Sheet or Certificate of Analysis?
A.
    
Supporting analytical information is reported in the Analytical Data Sheet (ADS) for each product.  Due to the sensitive and confidential nature of custom peptides, Peptides International does not make this information available online for download as with the catalog products.  Instead the ADS is included in each product's packaging.  Should you need more information on this, please contact us via email or phone.

Q.    How should a peptide be stored?
A.    Once a product arrives at your facilities, it should be stored at -20 °C or below.  The peptide should be stored with a desiccant to maintain a dry environment.

Q.    What salt form should I use for my custom peptide?
A.
    When ordering a custom peptide or a catalog peptide, the salt form of a peptide is important. In simple in vitro experiments, a trifluoroacetate (TFA) salt is usually acceptable. However, when experiments are performed in vivo or in cell culture, one may consider ordering an Acetate (-OAc) or hydrochloride (HCl) counter ion to minimize any issues associated with TFA. There is usually a small charge for exchange of the counter ion, but this is definitely worth it to eliminate some of the associated issues known for TFA especially in cell culture experiments and animal models.

Preparing Custom Peptide Solutions

Q.    How should a peptide solution be prepared?
A.    Since there is not a universal solvent for dissolving every peptide, this crucial step is not always straightforward as it may appear.  Because several solvent systems may be necessary until the desired conditions are achieved, always test a small sample of the peptide to determine the best solvents for complete solubilization.  When testing, begin with solvents that can be easily removed by lyophilization, such as water and acetic acid.  For this reason, it is not recommended to start with buffers which have high salt concentrations.

Determine the overall charge
Charged amino acids aid solubility in aqueous environments.  A sequence with little or no overall charge at any pH is not likely to be water soluble.

•    Hydrophobic amino acids:  Ala, Phe, Ile, Leu, Val, Pro, Met, Trp, Tyr, Cys
•    Positive Charges:  Lys, Arg, His and Free N-terminus
•    Negative Charges: Asp and Glu and Free C-terminus
First determine if the peptide is charged or neutral by calculating the overall charge of the peptide at pH 7.  Calculate overall charge by using the following values:
•    +1 for each basic residue (Lys, Arg, and N-terminus)
•    -1 for each acidic residue (Asp, Glu and C-terminus)
•    For His, use +1 at pH 6

If the overall charge of the peptide is negative then the peptide is acidic.  If it is positive then it is basic, and if zero then it is neutral.  

Charged peptides
For acidic peptides (and/or if the total number of charges of the peptide at pH 7 is greater than 25% of the total number of residues): use a small amount of 0.1M ammonium bicarbonate to dissolve the peptide, and then dilute it with water to the desired concentration.  Maintain pH around 7 and adjust pH as needed.
For basic peptides (and/or if the total number of charges of the peptide at pH 7 is between 10-25% of the total number of residues): use a small amount of 25% acetic acid to dissolve the peptide and dilute it with water to the desired concentration.  Adjust pH with 8 M NH4OH to desired pH for oxidation (4-7).  
For neutral peptides (and/or if the number of charges is greater than 25% of the total number of residues), use the strategy described for acidic peptides.  Otherwise, the use of organic solvents is recommended.  

Hydrophobic or neutral peptides
Hydrophobic peptides containing 50% to 75% hydrophobic residues may be insoluble or only partially soluble in aqueous solutions, even if the sequence contains 25% charged residues.  It is best to first dissolve these peptides in a minimal amount of stronger solvents such as acetonitrile, isopropyl alcohol, ethanol, and/or acetic acid, and then slowly add (drop wise) the solution to a stirred aqueous buffer solution.  If the resulting peptide solution begins to show turbidity, you might have reached the solubility limit and it will be futile to proceed.  Again, it is important to remember that the initial solvent of choice should be compatible with the experiment.
It is important to dissolve the peptide completely in the initial solvent (such as acetic acid, acetonitrile) because the rate of dissolution of peptides into these solvents is usually higher than in a water/solvent mixture.  If a water/solvent mixture is used first to dissolve the peptide, the final volume of solution may be larger than necessary.

Sonication
It may be necessary to sonicate the solution before determining if the solvent choice was appropriate.  Sonication should improve solubilization by breaking the solid peptide into smaller particles.  If the solution gels, becomes cloudy or turbid, or has visible particulates, the peptide has not dissolved completely but is suspended.  At this point, a stronger solvent is necessary and the solvents should be removed.  Begin again with the dry sample.

Liability Disclaimer
All products sold by Peptides International, Inc. are intended solely for laboratory and research use and should not be used in or on human subjects.  User assumes all risk of patent infringement by reason of use of material provided by Peptides International. Peptides International will not be responsible for damages arising from misuse of any product and is not responsible for the results of research using our products. 

 


Frequently Asked Questions about Peptides

 

Q.    AMC or MCA: Which one is it?
A.   
The fluorogenic compound, 7-Amino-4-MethylCoumarin (CAS Number: 6093-31-2, C10H9NO2), is often called AMC and is used in fluorogenic substrates for the detection of proteolytic enzyme activity. The release of AMC can be detected by fluorescence or absorbance. 1 Typically, the free compound is referred to as “AMC”. When it is bound to the C-terminus of an amino acid, it becomes 4-MethylCoumaryl-7-Amide, and Peptides International refers to the bound version as MCA, although it is sometimes still identified as “AMC”, even when bound. An example, is the compound, H-Lys-MCA, which is sometimes referred to as H-Lys-AMC.

1. C.P. Linn et al., Anal Biochem. 200(2), 400 (1992).

Q.    How do I find Assay Methods for my Enzyme Substrate?
A.   
The substrate assay methods can often be found in published literature or, if available, can be found here:

or feel free to contact us for more information.

Q.   What leads to different salt forms of peptides?
A.
   
Many catalog and custom peptides are present in the trifluoroacetate (TFA) salt form.  Peptides are often purified by reverse-phase High Performance Liquid Chromatography (rHPLC) using a TFA buffer.  The charge on a free amine in the peptide, either on the N-terminus or in a side chain, attracts the TFA counter ion.  In some cases, TFA salt is not the preference and an alternate salt form is preferred.  An additional step is done by using an ion exchange column to exchange the TFA salt with a preferred salt.  Some examples of alternative salt forms are acetate (-OAc) or hydrochloride (HCl).  Since converting from a TFA salt form to another salt form requires an additional step, there is often an additional small fee.


Frequently Asked Questions about Peptide Synthesis

Q.    What is a safer alternative to HOBt and 6-Cl-HOBt?
A.    OxymaPure® is an additive used in carbodiimide-mediated (DIC or EDC-HCl) reactions in the conversion of carboxylic acid to amide, with increased yield and decreased side reactions.   It is a safer and more effective alternative to HOBt and has no transport restrictions. Peptides International is an official distributor of OxymaPure®. Read more about OxymaPure® here. Other alternatives to HOBt and 6-Cl-HOBt are HCTU or HATU. View all peptide synthesis reagents here.

Q.    Where can I find more information on CLEARTM Resins?
A.    CLEAR (Cross-Linked Ethoxylate Acry- late Resins) was developed by George Barany and Maria Kempe at the University of Minnesota. These products retain the highly desirable solvation properties of polyethylene glycol (PEG) or of PEG-linked products but with greater convenience. Unlike resins for conventional liquid phase synthesis, CLEAR resin is a highly cross-linked solid support. It is produced in a bead form using a large-scale polymerization process developed at Peptides International. Read more about CLEAR resins here.

Q.    Do you carry a polymer-supported oxidant to help prepare disulfide bonds?
A.    CLEAR-OXTM, is a polymer-supported oxidant, which combines the power of solid phase chemistry with the versatility of solution-phase reactions. CLEAR-OXTM is a highly effective, polymer-supported reagent for the formation of disulfide-bonds.1,2 CLEAR resin is the polymer of choice due to its compatibility with both aqueous and organic environments. A lysine-preformed cyclic Ellman’s reagent [5,5’-dithiobis(2-nitrobenzoic acid) = DTNB] is covalently attached to a CLEAR polymeric support with a β-alanine spacer to yield CLEAR-OX.3,4Since the mechanism is based on peptide capture, sensitive residues such as Tyr, Trp, and Met are not affected, leading to increased purity and yield. These improved synthetic conditions allow for facile removal of the oxidant. Read more about CLEAR-OXTM and download resin conditioning and general methods here.

Q.    How do I choose an appropriate substitution for my resin of interest?
A.
    
Mesh is the number of square openings in a linear inch of screen or sieve. As the mesh number increases, the number of openings also increases while the size of the openings decreases. Therefore, larger mesh numbers correspond to smaller bead or particle sizes for a resin.

US Mesh

Inches

Microns

Millimeters

100

0.0059

149

0.149

120

0.0049

125

0.125

140

0.0041

105

0.105

170

0.0035

88

0.088

200

0.0029

74

0.074

230

0.0024

63

0.063

270

0.0021

53

0.053

325

0.0017

44

0.044

400

0.0015

37

0.037

 

Q.    What does the mesh number mean for a resin?
A.
    
The higher the substitution, the greater the points of attachment points there are. However, higher substitutions can lead to decreased synthesis efficiency and cause aggregation with peptides containing long or difficult sequences. This is caused by increased steric hindrance that accompanies high density of peptide chains present in the matrix. Lower substitutions are also better to use for cyclizing linear peptides.

Q.    The substitution for resins is shown in the units, meq/g. How do you convert this to mmol/g an appropriate substitution for my resin of interest?
A.
    
No conversion is required since meq/g is equivalent to mmol/g.


Do you have distributors?

Peptides International has a robust and valued network of distributors that allows an even more seamless dissemination of our products.  These partnerships include:

When ordering a custom peptide or a catalog peptide, the salt form of a peptide is important. In simple in vitro experiments, a trifluoroacetate (TFA) salt is usually acceptable. However, when experiments are performed in vivo or in cell culture, one may consider ordering an Acetate (-OAc) or hydrochloride (HCl) counter ion to minimize any issues associated with TFA. There is usually a small charge for exchange of the counter ion, but this is definitely worth it to eliminate some of the associated issues known for TFA especially in cell culture experiments and animal models.