Why molar concentration and not milligrams per millilitre
Two peptides at the same mass concentration are not at the same molecular concentration unless they happen to weigh the same. A milligram of a 500 dalton peptide contains twice as many molecules as a milligram of a 1,000 dalton one, and anything that depends on molecules meeting molecules, receptor occupancy, enzyme kinetics, stoichiometric binding, depends on the count.
That is why literature reports treatment concentrations in nanomolar or micromolar rather than in micrograms per millilitre. A published EC50 of 10 nM is a statement about molecules, and reproducing it means converting through molecular weight rather than matching the mass figure.
The three questions this calculator answers
Mass to molarity asks what concentration a known mass in a known volume produces. This is the everyday case: a vial reconstituted to a volume, expressed in molar terms.
Molarity to mass runs it backwards, and is the question you ask when planning: to make a given volume at a given molarity, how much material do you need to weigh out or withdraw. Dilution answers the third case, where you already have a stock and need a smaller working concentration from it.
The dilution relationship
C1V1 equals C2V2 is a statement of conservation: the amount of substance in the aliquot you take from the stock is the same amount that ends up in the diluted solution. Rearranging gives the stock volume needed, and the diluent volume is the difference between that and the final volume.
The relationship holds in any concentration units, as long as both concentrations are in the same ones. It does not need molecular weight at all, which is why the dilution tab has no molecular weight field.
Getting the molecular weight right
The molecular weight is the input most likely to be wrong, and an error in it passes straight through into the molar concentration. Three things commonly cause it.
First, the salt form: a peptide supplied as a TFA or acetate salt weighs more than the free base, and using the free base weight against a mass that includes counterions understates the molarity. Second, terminal modifications: an amidated C-terminus is 0.98 Da lighter than the free acid and an acetylated N-terminus is 42.01 Da heavier, so a weight calculated from the bare sequence can miss the catalogue value. Third, disulfide bridges, each of which removes 2.02 Da.
- •Use the weight that corresponds to the material you actually weighed or withdrew.
- •A catalogue weight that disagrees with a sequence calculation usually differs by a nameable modification, not by an error.
- •The molecular weight calculator computes the linear free-acid value; the modification mass calculator adds the terminal and post-translational changes.
Reading very small and very large numbers
Molar concentrations in peptide work span an enormous range, from millimolar stocks down to picomolar assay concentrations, so the calculator reports the same figure in molar, millimolar, micromolar and nanomolar simultaneously. Reading the one that puts the value between one and a thousand avoids most transcription errors.
How the molarity calculations work
Three rearrangements of one relationship, moles equals mass over molecular weight, with the units chosen so that the everyday case, milligrams in millilitres, needs no scaling factor at all.
mass to molarity: M = mass (mg) / MW (g/mol) / volume (mL) molarity to mass: mg = molarity (mM) x MW (g/mol) x volume (mL) / 1000 dilution: V1 = C2 x V2 / C1 diluent to add: Vd = V2 - V1
- Divide mass by molecular weight. Milligrams divided by grams per mole gives millimoles. This is the step that converts a quantity of material into a count of molecules.
- Divide by volume in millilitres. Millimoles per millilitre is moles per litre, which is molar. The units cancel so cleanly that no conversion factor appears in the everyday case.
- Scale for display. The molar figure is multiplied by 1,000, one million and one billion to give millimolar, micromolar and nanomolar, so the reader can pick the scale that reads naturally.
- Run it backwards for the planning case. Molarity times volume gives moles, and moles times molecular weight gives mass. The factor of 1,000 in the formula converts millimolar and millilitres into milligrams.
- Apply conservation for dilutions. C1V1 equals C2V2 rearranges to give the stock volume. The diluent volume is the final volume minus the stock volume, which the tool reports separately so you are not subtracting by hand.
What this method cannot tell you
- •It assumes the mass you enter is the peptide. For a salt form or a preparation below full purity, the actual peptide mass is lower and the true molarity is correspondingly lower.
- •It assumes complete dissolution. Undissolved material is not in solution and does not count towards concentration, however carefully it was weighed.
- •Volumes are treated as additive. Mixing a stock with a diluent gives a final volume very slightly different from the sum, which is negligible for aqueous solutions at these concentrations.
- •The dilution mode does not check that the stock volume it returns is physically measurable. A calculated 0.3 microlitre transfer is arithmetically correct and practically useless.
Peptide molarity calculator: frequently asked questions
The number of moles of a substance per litre of solution. A mole is a fixed count of molecules, so molarity is a concentration expressed in molecules rather than in mass.
One molar means one mole per litre. Peptide work usually lives several orders of magnitude below that, in millimolar stocks and micromolar or nanomolar working solutions.
Divide the mass by the molecular weight to get moles, then divide by the volume in litres. In practical units, milligrams divided by molecular weight divided by millilitres gives molar directly.
5 mg of a 1,419.5 Da peptide in 2 mL works out to 5 / 1419.5 / 2, which is 1.76 millimolar.
Because biological interactions count molecules. A receptor binds a molecule, not a milligram, so two peptides of different weight at the same mass concentration present different numbers of molecules to the same number of binding sites.
This is why published potency figures are given in molar terms. Reproducing a 10 nM condition means converting through molecular weight, not matching a mass concentration.
The one matching the material you actually have. A peptide supplied as a TFA salt weighs more than the free base, and using the free base weight against salt mass overstates how many molecules you have.
For a peptide with terminal modifications, the sequence-derived weight can also differ from the catalogue value: an amidated C-terminus is 0.98 Da lighter and an acetylated N-terminus 42.01 Da heavier.
It says the amount of substance is conserved. Whatever you take out of the stock ends up in the diluted solution, so concentration times volume before equals concentration times volume after.
Rearranged, the stock volume you need is C2 times V2 divided by C1. The diluent volume is the final volume minus that.
No. C1V1 equals C2V2 works in any concentration units as long as both sides use the same ones, so a dilution in mg/mL needs no molecular weight at all.
The molecular weight is only needed when you cross between mass and molar units.
Each is a thousandfold step down from the one before.
- •1 M = 1 mole per litre
- •1 mM = 0.001 M
- •1 µM = 0.001 mM
- •1 nM = 0.001 µM
The calculator shows all four at once so you can read the scale that puts your figure between one and a thousand, which is where transcription errors are least likely.
Multiply the molarity by the volume to get moles, then multiply by the molecular weight to get mass. The molarity-to-mass tab does this directly.
For 1 mM in 1 mL of a 1,419.5 Da peptide, that is 1.42 mg.
They are the free base weights for the named compounds and are accurate to within rounding at the last decimal place. They are not salt weights.
If your material is a salt, add the counterion mass before using the figure. The salt form converter calculates that from the number of counterions.
Because they are two different molecules. GHK is the free tripeptide Gly-His-Lys at 340.38 Da. GHK-Cu is that peptide complexed with a copper ion, at 403.93 Da.
Both are in the preset list under their own names. Using the copper complex's weight for the free peptide, or the reverse, is roughly an 18 percent error.
Yes, directly. If a nominal 5 mg is only 90 percent peptide, then 4.5 mg of peptide is present and the molarity is 10 percent lower than the calculation from the label suggests.
For work where the absolute concentration matters, correct the mass first with the net peptide content calculator and enter the corrected figure here.
Undissolved material is not in solution and contributes nothing to concentration, so the real molarity is lower than the calculation, by an amount you cannot see.
This is the failure mode behind assay results that are inexplicably weak. The solubility predictor flags sequences likely to need a co-solvent before you commit material to plain water.
Yes. The arithmetic is identical, and molecular weight is just a larger number. Enter the protein's weight in daltons in place of a peptide's.
For very large proteins the molar concentrations get small quickly: 1 mg/mL of a 66,000 Da protein is only about 15 micromolar.
Because the stock is too concentrated for the dilution you are asking for in a single step. A 1,000-fold dilution into 1 mL calls for a 1 microlitre transfer, which most pipettes cannot deliver reproducibly.
The fix is a serial dilution: two 30-fold steps are far more accurate than one 1,000-fold step. The serial dilution calculator lays out the scheme.
Slightly, because volume changes with temperature and molarity is defined per litre of solution. Between room temperature and refrigeration the effect is a fraction of a percent.
Molality, which is per kilogram of solvent, avoids this entirely. It is rarely used in peptide work because the effect is too small to matter.
Arithmetically exact. The practical precision is set by your inputs: the accuracy of the balance or the syringe, whether the material fully dissolved, and whether the molecular weight matches what you actually have.
Of those, the molecular weight is the one most often wrong and the easiest to check.
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