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Sidechain Tyrosine Resin Loading

Applicable for sequences bound for on-resin cyclization

Overall Reaction

This method describes the loading of tyrosine through the side chain for the purposes of on-resin cyclization. This chemistry allows for on-resin cyclization and global methylation. It is, however, not stable to the reducing conditions used for either NBS or TFA-mediated in-sequence selective methylation. In contrast to resin bound for solution-phase cyclization, this resin is not loaded to maximum capacity; instead, to prevent dimerization during cyclization, it is loaded no higher than 0.45 mmol/g. This method was adopted from Arthur Rand and Rushia Turner.1

This page assumes the loading of 5 grams of 2-chlorotrityl resin.

Step 1: Tyrosine Preparation

  1. Take an oven-dried 250 mL round-bottom flask and cool it under a stream of nitrogen.
  2. Add 2.25 mmol (0.99 g) of Fmoc-O-Allyl-Tyrosine to the flask (previously prepared or purchased).
  3. Rubberband a Kimwipe on top and vacuum desiccate overnight.

Step 2: Loading

  1. Acquire ~100 mL of dry DCM from the solvent purification system.
  2. Add ~75 mL of DCM (via syringe or cannula) to the flask charged with tyrosine.
  3. Add 4 eq (1.17 mL) of sieve-dried DIPEA.
  4. Dissolve as best you can.
  5. Add 5 g of unloaded 2-chlorotrityl resin.
  6. Add a head of nitrogen (blowing away the white smoke that forms).
  7. Cap with a septum and a nitrogen balloon.

  8. Shake overnight on the orbital shaker.

    Tip

    Slowly rolling the flask will collect stray resin that collects above the solvent line.

Example Reaction Table

Name g MW mmol eq d mL
2-Cl-Trityl resin 5 n/a n/a n/a n/a n/a
Fmoc-OAllyl-Tyrosine 1 443.41 2.25 1 n/a n/a
DIPEA 1.2 129 9 5 0.74 1.6

Step 3: Capping

Collecting Resin

  1. Collect the resin into a solid-phase synthesis vessel using DCM and a funnel.
  2. Wash 3x with DCM.

Prepare Capping Solution

Make a 17:2:1 solution of DCM:MeOH:DIPEA. For this scale, 51 mL of DCM, 6 mL of MeOH, and 3 mL of DIPEA works well.

Capping

  1. Add 20 mL of capping solution, mix briefly, drain.
  2. Add 20 mL and shake for 20 minutes.

  3. Add the final 20 mL and shake for another 20 minutes.

    Tip

    Make sure to release the pressure from the synthesis vial before shaking or the cap will eject.

  4. Wash 3x DMF.

  5. Wash 3x DCM.
  6. Put a rubberbanded Kimwipe on top and pull air over the resin overnight for future loading determination.

Bibliography

  1. Rand et al., Int J Pept Res Ther 2012

  1. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3488302/#!po=79.5455 ↩