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Pictet-Spengler route to Praziquantel
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3rd January 2012 @ 23:11
Extension of the procedure performed here. The previously performed experiment resulted in a mixture of the enediamide intermediate and the starting material (as indicated by proton NMR). The following experiment was performed in an attempt to drive the reaction to completion.

Stoichiometric acid mediated Pictet-Spengler (PS) cyclisation of the Ugi-intermediates (KAB5-1 and KAB6-1) to give racemic PZQ (KAB3-4) and the N-benzoyl PZQ analogue (KAB7-2).

products.png


Scheme
Scheme%20Round%202.png
table.jpg

Hazard and Risk Assessment
As for KAB3-3 and KAB7-1 (here).

Relevant Experiments
Preparation of the PZQ analogue Ugi-intermediate (KAB5-1)
Preparation of the N-benzoyl PZQ analogue Ugi-intermediate (KAB6-1)
Acid-mediated PS cyclisation to give racemic PZQ (KAB3-3), the dimethoxy N-benzoyl PZQ analogue (KAB8-1), the dimethoxy PZQ analogue (KAB1-2) and the N-benzoyl PZQ analogue (KAB7-1)
Stoichiometric acid-mediated Pictet-Spengler of MW29-4 to give racemic PZQ (KAB3-1 and KAB3-2)


Procedure

04/01/12

The starting materials for the reaction were added to 100 mL round bottom flasks then dissolved in toluene.

KAB6-1 in toluene before the addition of MSA
KAB5-1 in toluene before the addition of MSA


Following the addition of methanesulfonic acid (MSA), the reaction mixtures were reflux heated at 70C.

Reaction Setup

Aliquots (0.1 mL) were extracted from the reaction mixtures over the duration of the reflux. The aliquots were quenched in saturated sodium bicarbonate (2 mL) and EtOAc (1 mL). The reaction progress was monitored by TLC of the aliquots against the corresponding starting materials.

KAB3-4:
Starting material (KAB5-1) added - 1.0 g, 2.5 mmol.
MSA added at 09:45 - 0.4 mL, 2.8 mmol
Toluene - 10 mL, ~50 eq.
1st Aliquot - 09:53 (8 min), T = 37 C --> Pale yellow solution
2nd Aliquot - 10:00 (15 min), T = 60 C
3rd Aliquot - 10:05 (20 min), T = 68 C
10:07 (22 min) --> Reached 70 C
10:14 (29 min) --> Dark red colour
4th Aliquot - 10:15 (30 min), T = 73 C
5th Aliquot - 10:33 (48 min), T = 71 C
6th Aliquot - 10:46 (1 h 1 min), T = 71 C
7th Aliquot - 11:39 (1 h 53 min), T = 71 C
8th Aliquot - 12:09 (2 h 24 min), T = 70 C
9th Aliquot - 13:13 (3 h, 28 min), T = 70 C
10th Aliquot - 13:58 (4 h, 13 min), T = 71 C
11th Aliquot - 14:48 (5 h 03 min), T = 70 C
12th Aliquot - 15:42 (5 h 57 min), T = 70 C
13th Aliquot - 16:49 (7 h 4 min), T = 70 C
14th Aliquot - 08:01 (22 h 16 min), T = 71 C
15th Aliquot - 14:33 (1 d 4 h 48 min), T = 70 C
16:07 (1 day 6 h 22 min) --> turned up controller setpoint to 110 C, see KAB3-1
16th Aliquot - (1 d 7 h 17 min), T = 109 C
17th Aliquot - (1 d 22 h 22 min), T = 110 C
18th Aliquot -

KAB7-1
Starting material (KAB6-1) added - 1.1 g, 2.8 mmol
MSA added at 09:47- 0.4 mL, 2.8 mmol
Toluene - 11 mL, ~50 eq.
1st Aliquot - 09:55 (8 min), T = 58 C
2nd Aliquot - 10:01 (14 min), T = 70 C --> Reached 70 C, dark red colour
3rd Aliquot - 10:07 (20 min), T = 74 C
4th Aliquot - 10:17 (30 min), T = 74 C
5th Aliquot - 10:34 (47 min), T = 79 C
10:45 (58 min), T = 86 C --> manually turned down heat.
6th Aliquot - 10:47 (1 h 0 min), T = 78 C
11:06 (1 h 19 min), T = 66 C --> manually turned up heat.
7th Aliquot - 11:39 (1 h 52 min), T = 67 C
8th Aliquot - 12:11 (2 h 24 min), T = 70 C
9th Aliquot - 13:14 (3 h 27 min), T = 73 C
10th Aliquot - 13:59 (4 h 12 min), T = 73 C
11th Aliquot - 14:49 (5 h 2 min), T = 71
12th Aliquot - 15:44 (5 h 57 min), T = 70 C
13th Aliquot - 16:51 (7 h 4 min), T = 65
14th Aliquot - 08:03 (22 h 16 min), T = 64
15th Aliquot - 14:35 (1 d 4 h 48 min), T = 72
16:09 (1 day 6 h 22 min) --> turned up controller setpoint to 110 C, see KAB3-1
16th Aliquot - (1 d 7 h 19 min), T = 128 C
17th Aliquot - (1 d 22 h 21 min), T = 108 C
18th Aliquot - (2 d 2 h 52 min), T = 110 C
Take off heat - (2 d 3 h 29 min)

Monitoring the reaction by TLC
All TLCs were performed in EtOAc/hexane, 3:1, v/v with a KMnO4 stain.
TLC of reaction mixtures (1)
TLC of reaction mixtures (2)
TLC of reaction mixtures (3)
TLC of reaction mixtures (4)

Note: did not mark TLC plates 3 or 4 for the KAB3-4 reaction mixture. "Fragmentation spots" were weak and faded upon heating. The images were captured before the plate was completely dry and annotation was avoided to prevent scratching of the stationary silica phase.

05/01/12 - Continued monitoring of the reaction by TLC
TLC of reaction mixtures (5)
TLC of reaction mixtures (6)
TLC of reaction mixtures (7)


Recorded the mass spectra (MS) of the 12th and 14th collected aliquots for both reaction mixtures.

MS KAB7-2, Aliquot 14

Peaks of Interest, m/z(ra%): 391.07(100), 967.80(94), 633.20(53), 825.20(469), 665.20(40), 413.07(40), 294.93(44).

MS KAB7-2, Aliquot 12

Peaks of Interest, m/z(ra%):

MS KAB3-4, Aliquot 14

Peaks of Interest, m/z(ra%):

MS KAB3-4, Aliquot 12

Peaks of Interest, m/z(ra%):

The MS of all four samples showed no identifiable product peaks. Despite being crude, the two aliquots of both samples contained the same peaks as those in MNR14-1 and MNR12-1 (the MS of which were obtained at the same time).

NOTE: MNR14-1 and MNR12-1 were prepared by a similar procedure X ago, the expected structure of which are identical to KAB3-4 and KAB7-2 (i.e. PZQ and the N-benzoyl PZQ analogue).

Expected MS Peaks
expected%20MS.png
expected%20mass.jpg

The temperature of the reaction mixture was increased to 110 C to encourage product formation (the TLCs showed no further increase in intensity of the desired spot from 7 hours to ~30 hours).

After increasing temperature of reaction mixture to 110 C:
KAB7-2 reaction mixture following increase to 110 C
TLC of reaction mixtures (8)

NOTE: KAB7-2 reaction mixture spiked at 138 C (despite setpoint of controller of 113 C). Holding 107-111 C, time = 1 d 8 h after reaction start.


06/01/12 - Continued monitoring of the reaction by TLC

TLC of reaction mixtures (9)


KAB3-4 was removed from heat 51 hours after the addition of MSA. Following dilution with dichloromethane (DCM, 30 mL), the reaction mixture was quenched with saturated NaHCO3 (50 mL). The organic DCM fraction was separated and the aqueous layer further extracted with DCM (3 x 30 mL).

KAB3-4 after removal from heat
Dilution of the reaction mixture with DCM
KAB3-4 first extraction with DCM
KAB3-4 second extraction with DCM
KAB3-4 third extraction with DCM
KAB3-4 aqueous fraction


The DCM fractions were combined then washed with saturated NaCl solution (50 mL), dried over MgSO4 and filtered. The solvent was removed under reduced pressure to give crude KAB3-4 as a highly viscous, dark brown oil (412 mg).

Separating KAB3-5 following washing combined DCM layers with NaCl soln

NOTE: typo in above picture description. Label "KAB3-5" should read "KAB3-4"

KAB7-2 was removed from heat 51 hours after the addition of MSA. The reaction mixture was worked up identically to KAB3-4 to give crude KAB7-2 as a dark brown, highly viscous oil (465.3 mg).

KAB7-2 after removal from heat
Extracting the aqueous fraction of KAB7-2
KAB7-2 second extraction with DCM
KAB7-2 aqueous fraction
Filtering KAB7-2 after drying


TLC of concentrated crude KAB3-4 and KAB7-2
TLC of crude KAB3-4 and KAB7-2


09/01/12
Both crude samples solidified over the weekend.


23/01/12
The reaction was unsuccessful for the cyclisation to PZQ, and inconclusive for the N-benzoyl analogue. Based on the odour and the TLC of the final crude products, it is likely that hydrolysis of the starting material or the enediamide intermediates occurred to give PZQamine or some tautomer of it.

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NOTES:

- Smell of isonitrile when extracting first aliquots of both reaction mixtures.

- Trouble with hotplate/controller of KAB7-2. At 60 min, temp was at 86.6 C. Had to manually turn down temperature. Then it dropped to 66 C at 82 min. Again, fixed it manually. Temp fluctuating between 67-70 up to now (time is 1h 47min).

- Regarding second TLC of KAB3-4: I almost left it too long, but I managed to catch it just before the mobile phase went off the edge. On heating, just after staining the plate, the slow spots started to appear in all spotted fractions (i.e. 3-6), with decreasing intensity. Some shadows of the same spots, possibly with relatively similar Rf appeared on the analogous fractions of KAB7-2. I attempted to resolve them by gently heating, but managed to overheat it, making them disappear completely. The area that they appeared is indicated by the arrow drawn on the plate.

- KAB7-2 went red to dark red within ~20 min. I paused stirring momentarily, and the reaction mixture looked to be two phases, with a yellow top layer and a deep red bottom layer, very similar to that observed in preparation of KAB3-1 and KAB3-2. Ethoxy leaving group, possibly containing MSA-ethoxy adduct?

- The 3rd TLC plate was not marked with a pencil. Started to, but it was still damp, and the silica was scratched off. Took picture instead, to try and record fragment spots before they faded. Will re-do...done.

- Getting something with the same Rf as expected product for KAB3-4 (i.e. PZQ). Unsure whether it is PZQ, or other fragment. See TLC 3 and 4.

- The product KAB7-1 started to appear after ~2.5 hours. Or not. Not sure what was appearing, possibly contamination from capillary?

- At ~26 hours, both solutions appear almost black.

- KAB7-2 mixture started splattering. Black deposits on the side of the flask.

- From the TLC of aliquot 15. The spot on the baseline is getting more intense.

- KAB3-5 was worked up before KAB3-4. The tar of KAB3-5 did not dissolve in EtOAc, but was easily dissolved in DCM. Consequently, the work-up of KAB3-4 was done in DCM and no EtOAc.

- There was a lot of debris in the KAB7-2 reaction mixture. The tar-like substance, presumably organic decomposition products were sticky and difficult to remove from the glassware. The substance is clearly and visibly stuck to the separating funnel in the pictures posted. Increased proportion of the tar is possibly due to the high temperature spikes of the reaction mixture (up to 138 C). The substance was similar to the "sludge" in the bottom of all Pictet-Spengler cyclisation reaction mixtures (with the prefix KAB) after removal from heat. Unlike the tar in KAB3-4 and KAB3-5 (the reactions of which were carried out simultaneously), it did not readily dissolve in DCM.
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Links
More photos here
Linked Posts
Attached Files
Scheme KAB3-4 and KAB7-2
Table
Structures
TLC of reaction mixtures (1)
KAB5-1 in toluene before the addition of MSA
KAB6-1 in toluene before the addition of MSA
Reaction Setup
TLC of reaction mixtures (2)
TLC of reaction mixtures (3)
TLC of reaction mixtures (4)
TLC of reaction mixtures (5)
TLC of reaction mixtures (6)
TLC of reaction mixtures (7)
TLC of reaction mixtures (8)
KAB7-2 reaction mixture following increase to 110 C
MS KAB3-4, Aliquot 12
MS KAB3-4, Aliquot 14
MS KAB7-2, Aliquot 12
MS KAB7-2, Aliquot 14
TLC of reaction mixtures (9)
Reaction mixture after removal from heat
KAB3-4 after removal from heat
KAB3-4 first extraction with DCM
KAB3-4 second extraction with DCM
KAB3-4 third extraction with DCM
Separating KAB3-5 following washing combined DCM layers with NaCl soln
Dilution of the reaction mixture with DCM
TLC of crude KAB3-4 and KAB7-2
KAB7-2 after removal from heat
Extracting the aqueous fraction of KAB7-2
KAB7-2 second extraction with DCM
KAB3-4 aqueous fraction
KAB7-2 aqueous fraction
Filtering KAB7-2 after drying
Expected MS
expected MS structures
Comments
Re: Extended reaction time: Acid-mediated Pictet-Spengler reaction to give PZQ (KAB3-4) and the N-benzoyl PZQ analogue by Matthew Todd
4th January 2012 @ 00:50
TLCs - good to have the S/M, but you really also need PZQ for reference. It's possible you're making the enediamide, which would appear to be a stable, discrete spot on the plate but which would have a very different Rf value to PZQ (or benzoyl analog).

Also - your URLs are becoming rather lengthy. Any way you could shorten the post title to something informative, then include a subheading that gives the reason for the experiment but which does not then become part of the link?

Oh, and Happy New Year. Great you're in the lab. I'm on holiday, as you can tell.
Re: Extended reaction time: Acid-mediated Pictet-Spengler reaction to give PZQ (KAB3-4) and the N-benzoyl PZQ analogue by Katrina Badiola
4th January 2012 @ 01:48
Started including the PZQ out of the bottle from the 3rd TLC. Very different Rf to the enediamide (which we discovered last time), but has a similar Rf to the starting material.
How do I do a subheading - or would it just be the first sentence like the text in the post.