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5th January 2012 @ 00:09
********Experiment still in progress with regular updates. Last updated 13/01/12

Modification of initial method (here).

Scheme
Scheme%20KAB3-5.png
Fixed%20Table.jpg

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

Following Experiments
Reattempt acid-mediated Pictet-Spengler reaction to give PZQ (KAB3-6)

Procedure

KAB5-1 (0.420 g) was dissolved in toluene (4 mL) and added portionwise to methanesulfonic acid (MSA, 150 uL) at 0 C. The reaction mixture was reflux heated at 70 C and the progress monitored by TLC.

MSA added + Start heating @ 10:41
Reached 70 C @ 10:54 (+13 min)

Reaction mixture at 21 min


Aliquots (10-30 uL) were extracted over the duration of the reflux. The aliquots were quenched in saturated sodium bicarbonate (1 mL) and EtOAc (0.5 mL). The reaction progress was monitored by TLC of the aliquots against the starting material KAB5-1, the expected product, PZQ and analogous fractions collected from the KAB3-4 reaction (EtOAc/hexane, 3:1, v/v; KMnO4 stain).

Aliquot 1 - 10:50 (9 min), T = 45 C
Aliquot 2 - 10:54 (13 min), T = 70 C
Aliquot 3 - 11:03 (22 min), T = 72 C
Aliquot 4 - 11:21 (40 min), T = 71 C
Aliquot 5 - 11:56 (1 h 15 min), T = 70 C
Aliquot 6 - 13:58 (3 h 17 min), T = 70 C
Aliquot 7 - 14:35 (3 h 54 min), T = 70 C
16:16 --> turned up controller set point to 110 C, see KAB3-1
Aliquot 8 - 16:45 (6 h 4 min), T = 99 C
Aliquot 9 - 08:16 (21 h 35 min), T = 110 C
Aliquot 10 - 10:15 (23 h 24 min), T = 110 C
10:30 --> taken off heat.
TOTAL REFLUX TIME: 23 h 49 min.


Monitoring the reaction by TLC
TLC of KAB3-5, 13-22 min

From L to R:

TLC of KAB3-5, 22 min - 4 h

Fastest spot above aliquots 6 and 7 were visible under long wave UV. Didn't resolve well with KMnO4 stain.

Recorded mass spectrum of extracted aliquot 6:
MS of Aliquot 6 (1)
MS of Aliquot 6 (2)
MS of Aliquot 6 (3)

WRITE COMMENT OF MASS SPEC HERE AND WHY YOU MSed IN THE FIRST PLACE
Note on the MS: the sample analysed was very dilute. Posted three different averages of the same spectrum. There were also multiple injections of the sample in the one spectra.

TLC of KAB3-5, 4-6 h

6 = Aliquot 7; 7 = Aliquot 8. Top spots above aliquots 7 and 8 were visible on heating, before fading. No longer visible under long wave UV.


06/01/12 - Continued monitoring the reaction by TLC
Reaction mixture at 24h
TLC of KAB3-5, 6-22 h



The TLC of the extracted aliquot 9 indicated consumption of the enediamide intermediate. Faint spots were visible with similar Rf to PZQ. The reaction mixture was removed from heat and allowed to cool, ~24h after the addition of MSA.

TLC of KAB3-5, 4-24 h
Reaction mixture after removal from heat


The reaction mixture was diluted with EtOAc (25 mL), transferred to a separating funnel, then quenched with saturated NaHCO3 (25 mL). The aqueous layer was extracted with EtOAc (2 x 25 mL) and the EtOAc fractions combined.

Dilution with EtOAc
Reaction mixture flask after dilution with EtOAc and transfer to separating funnel
Third extraction with EtOAc


The aqueous layer was then extracted with dichloromethane (DCM, 3 x 25 mL). The DCM fractions were combined then washed with saturated NaCl solution (25 mL).

Reaction mixture flask following rinse with DCM
First extraction with DCM
Second extraction with DCM
Third extraction with DCM
Aqueous KAB3-5 fraction


Both the combined EtOAc fractions and DCM fractions were dried over MgSO4, filtered and then combined. The dark solution was concentrated under reduced pressure to give crude KAB3-5 as a dark reddish-brown, highly viscous oil (180 mg).

Final crude KAB3-5
TLC of crude KAB3-5

TLC from L to R : (1)Crude KAB3-4 ;(2) Crude KAB3-5;(3) Praziquantel;(4) Crude KAB7-2;(5) Previously prepared PZQ amine in CDCl3


09/01/12
Amount Loaded = 54.4141 - 54.2610 = 153 mg

Purification by silica gel column chromatography in MeOH/EtOAc, 0-100%, v/v. Collected ~60 fractions.

Columning crude KAB3-5


Flask Weight:
Fraction 1: Flask = 21.3259 g (Flask + Fraction = 21.3377 g) - 11.8 mg
H-NMR of KAB3-5 fraction 1 (raw data)


Fraction 2: Flask = 20.3931 g (Flask + Fraction = 20.4004 g) - Yield: 7.3 mg
H-NMR of KAB3-5 fraction 2 (raw data)


Fraction 3: Flask = 20.8333 (Flask + Fraction = 20.8414 g) - Yield: 8.1 mg
H-NMR of KAB3-5 fraction 3 (raw data)


Fraction 4-5: Flask = 61.828 g (Flask + Fraction = 61.879 g) - Yield: 5.1 mg
H-NMR of KAB3-5 fractions 4-5 (raw data)


Fraction 6-8: Flask = 12.4963 g (Flask + Fraction = 12.5107 g) - Yield: 14.4 mg

Fraction 9-13 - Yield?

Fraction 14-17 - Yield: 5.7 mg

Fraction 18-24: Flask = 16.242 g (Flask + Fraction = 16.2500 g) - Yield: 8.0 mg

Fraction 25-31 - Yield?
H-NMR of KAB3-5 fractions 25-31 (raw data)


Fraction 32-44 - Yield: 13.8 mg

Fraction 45-57 - Yield: 18 mg

Final Fraction: Flask = 21.3312 g (Flask + Fraction = 21.3597 g) - Yield: 28.5 mg

Approximate Total Recovery: 120 mg, 78%.


Conclusion (written 23/01/12)
The experiment was unsuccessful. The final product is likely a mixture of decomposition products. Complete consumption/reaction of the enediamide intermediate occurred some time between extraction of aliquot 7 and 9.
The recovered fractions from the attempted purification contained various impure fragments.


Related Experiments
Acid-mediated Pictet-Spengler reaction to give PZQ (KAB3-4) and the N-benzoyl PZQ analogue (KAB7-2)
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)
Acid-mediated Pictet-Spengler of MW29 to give rac-PZQ MNR13-1



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NOTES 05/01/12:
- The reaction mixture went dark red much quicker (<20 min) than KAB3-4 did. Heating was steady up to 70 C.
- I wonder if there's a way to annotate attached photos in here.

NOTES 06/01/12:
RE: The first TLC performed this morning, i.e. containing aliquots 7, 8, 9 and comparison to PZQ and KAB3-4 (aliquot 17). The intensity of the spots was very low. Required a bit more heating than usual. Unsure whether product has degraded overnight, or the sample was too dilute (despite multiple spots). Latter is more likely, as very faint spots appeared, corresponding to enediamide and something with the same Rf as expected product, PZQ (unless the majority of sample has degraded into the yellow dot stuck at the base line).

- FLASK OF CRUDE: 54.2610 g

- The concentrated crude product had a very strong, sweet smell. Possibly the PZQ amine --> finally hydrolysed that amide. I was wondering why that hadn't occurred before under acidic, reflux conditions. The reaction may work if the reflux temperature is set to 110C from the start. I don't know what the reaction did between 6 and 21 hours after the addition of MSA. Possibly went to completion in that time frame.
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Linked Posts
Attached Files
Scheme KAB3-5
Table KAB3-5
TLC of KAB3-5, 13-22 min
Reaction mixture at 21 min
TLC of KAB3-5, 22 min - 4 h
TLC of KAB3-5, 4-6 h
TLC of KAB3-5, 6-22 h
Reaction mixture at 24h
MS of Aliquot 6 (1)
MS of Aliquot 6 (2)
MS of Aliquot 6 (3)
Final crude KAB3-5
TLC of KAB3-5, 4-24 h
Reaction mixture flask after dilution with EtOAc and transfer to separating funnel
Combined DCM and EtOAc fractions
Third extraction with DCM
Dilution with EtOAc
Reaction mixture after removal from heat
Third extraction with EtOAc
Reaction mixture flask following rinse with DCM
First extraction with DCM
Second extraction with DCM
Aqueous KAB3-5 fraction
TLC of crude KAB3-5
Table KAB3-5 (fixed)
Columning crude KAB3-5
H-NMR of KAB3-5 fraction 1 (raw data)
H-NMR of KAB3-5 fraction 2 (raw data)
H-NMR of KAB3-5 fraction 3 (raw data)
H-NMR of KAB3-5 fractions 4-5 (raw data)
H-NMR of KAB3-5 fractions 25-31 (raw data)