Modification and Functionalization of the Guanidine Group by Tailor-made Precursors

The overall goal of this procedure is to provide a convenient synthetic access to guanidine functionalized peptides. This method can help answer key questions in the field of muscle peptide chemistry such as the development of integrin ligands or GPCR ligands. The main advantage of this technique is that it allows an easy, synthetic access to functionalized guanidines, completely compatible with SPPS.

The technology will be especially valuable for tactile molecules because the functional groups, which are modified, are typically deteriorating biomolecules. In our case, we aim for molecules which can be used fpr molecular imaging of distinct properties of individual cancers and for their specific treatment;an important issue for personalized medicine. To begin this specific procedure, add one gram of boc-pyrazol carboxamaidine and one gram of PPh3 to a 50 milliliter round-bottomed flask.

Subsequently, add 10 milliliters of dried THF to a 50 milliliter round-bottomed flask in an inert atmosphere via a rubber septum. Using a syringe, add 194 microliters of dry methanol. Stir the solution at room temperature.

Using a syringe, add a prepared solution of diisopropyl azodicarboxylate in THF dropwise over 15 minutes. Then, let the solution stir for two hours at room temperature. Using a rotary evaporator, remove the solvent under reduced pressure.

After this, dissolve the crude product in one milliliter of DCM. Next, let a let a flash column with 100 grams of silica, in 10%ethyl acetate in a pentane solution. Load the dissolved crude product on the column and add the the solvent under pressure.

Collect, identify and combine the fractions as outlined in the text protocol. Then, using a rotary evaporator, evaporate the solvent under reduced pressure to obtain the desired compound as a yellow, viscous oil. First, add one gram of boc-pyrazole carboxamidine, one gram of PPH3 and 0.9 grams of Dde-6-Aminohexanol to a 50 milliliter round-bottomed flask.

Then, add 10 milliliters of dry THF in an inert atmosphere using a rubber septum. Afterwards, dissolve the starting materials at room temperature. Using a syringe, add a prepared solution of diisopropyl azodicarboxylate in THF dropwise over 15 minutes.

Stir the solution for two hours at room temperature. After this, use a rotary evaporator to remove the solvent under reduced pressure. Dissolve the crude product in 1 milliliter of DCM.

Next, load a flash column with 100 grams of silica in a two to one solution of pentane and ethyl acetate. Load the dissolved crude product on the column and add the solvent under pressure. Collect, identify and combine the fractions as outlined in the text protocol.

Using a rotary evaporator, evaporate the solvent under reduced pressure to obtain the desired product. To begin, dissolve 1.4 grams of S-methylisothiourea and 2.2 grams of boc-anhydride in a vigorously-stirring biphasic solution of DCM and saturated aqueous sodium bicarbonate. Let the mixture stir for 24 hours at room temperature.

After this, pour the mixture into a separatory funnel. Separate the layers and extract the aqueous phase three times with DCM. Then, combine the organic phases.

Dry the combined organic phases with sodium sulfate. After removing the solvent using a rotary evaporator, dissolve the crude product in 2 milliliters of hexane. Load a flash column with 100 grams of silica in a four to one solution of hexane and ethyl acetate.

After this, load the dissolved crude product on the column and add the solvent under pressure. After collecting and identifying the fractions, combine the desired fractions. Using a rotary evaporator, evaporate the solvent under reduced pressure to obtain 1.1 grams of Boc-S-methylisothiourea.

Dissolve 250 milligrams of the collected Boc-S-methylisothiourea in 10 milliliters of dried DMF in a 50 milliliter round-bottomed flask in an inert atmosphere at room temperature. Use a syringe to add 440 microliters of DIPEA. Using a syringe, add 245 microliters of acetic anhydride dropwise while stirring.

Let the mixture stir for one hour at room temperature. Then, use a syringe to add two milliliters of methanol and stir for 15 minutes. After removing the solvent under reduced pressure, using a rotary evaporator, dissolve the crude product in one milliliter of DCM.

Load a flash column with 60 grams of silica in a three to one solution of hexane and ethyl acetate. Next, load the dissolved crude product and add the solvent under pressure. Collect, identify and combine the fractions as outlined in the text protocol.

Using a rotary evaporator, evaporate the solvent under reduced pressure to obtain 210 milligrams of the desired product as a white solid. To begin, dissolve a small amount of the orthogonally deprotected cyclic peptide in a small volume of DMF. Add two equivalents of DIPEA.

And two equivalents of the guanidinylation precursor. Then, let the solution stir for two hours at room temperature. Once the reaction is complete, remove the solvent.

Re-dissolve the guanidinylated cyclic peptide in acetyl nitrile and perform semi-preparative HPLC purification as outlined in the text protocol. After this, in a small glass vile, add a prepared solution of trifluoroacetic acid, water and triisopropylsilane to the purified product. Stir this mixture for one hour at room temperature, and then observe the complete deprotection with ESIMS.

Add 10 milliliters of ice-cold ether to a 50 milliliters centrifuge tube. Using a Pasteur pipette, add the deprotected peptide dropwise to the ether. Centrifuge the suspension at 5, 000 times g for five minutes.

After this, wash the pellet with ice-cold ether. In centrifuge at 5, 000 times g for five minutes. Repeat this washing and centrifugation once more.

Then, dissolve the product in 2 milliliters of HBLC-grade water and purify the product using semi-preparative HPLC as outlined in the text protocol. In this study, an easy method in the modification of the guanidine group in peptidic glygins is presented. The reactions'progress is monitored using high-pressure liquid chromatography.

For larger residues on the guanidine group, two hours is often not enough time for the reaction to reach completion. Thus, the reaction is continued and the compounds are purified by semi-preparative HPLC. A small amount is then diluted with DMSO to obtain a stem solution for the biological evaluation in an al-ee-so-like solid phase binding assay.

All the compounds analyzed show a relatively high affinity of the integrant subtype alpha v beta three, And high selectivity against the integrant subtype alpha five beta one. We first had the idea for this method when we were looking for a standard method for guanidinylation during SPPS. After watching this video, you should have a good understanding of how to prepare tailor made guanidinylation precursors, in order to modify or functionalize the guanidine group of your target peptide.