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24th November 2011 @ 22:31
The modified Hoffman procedure for the preparation of (2-isocyanoethyl)benzene for later use in the Pictet-Spengler cyclisation to PZQ and the N-benzoyl analogue.

Scheme%204-1.png

KAB4-1%20table.tjpg

Risk Assessment
Risk Assessment


01/12/11

Procedure
Sodium hydroxide pellets (60 g) were gradually added to water (60 mL) in a 500 mL round bottom flask. A solution of benzyltriethylammonium chloride (400 mg, 1.76 mmol) in dichloromethane (DCM) (60 mL), chloroform (16 mL, 200 mmol) and 2-phenylethylamine (24.3 mL, 200 mmol) in a pressure equalising dropping funnel was added gradually over 12 minutes. The reaction mixture was heated at reflux (38-42 C) for 4 hours.

Reflux Timeline:
1:45 pm: The reaction mixture reached reflux temperature of ~40 C.
3:15 pm: Formation of a light brown froth-like-solid, sticking to the sides of the flask. Stirrer turned up. Reflux temperature fluctuating between 34-42 C.
3:30 pm: Reaction frothing more vigorously.
4:05 pm: Reaction mixture a darker brown, froth/solid light brown. Oil bath temperature holding steady around 40.0-40.6 C.
5:10 pm: More brown precipitate in reaction mixture.

5:45 pm: The reaction mixture was removed from the oil bath, diluted with ~50 mL of ice and water then transferred to a separating funnel before removal of some of the organic layer. The solution consisted of a brown emulsion and the organic-aqueous interface was not clear. Water (50 mL) and DCM (100 mL) was added to the solution and the emulsion began to dissolve. After settling for 5 min, the organic fraction was removed, the aqueous layer extracted with DCM (2 x 50 mL) and the organic fractions combined. The organic fractions were washed with hydrochloric acid (1 M, 3 x 100 mL) followed by brine (100 mL). After drying over magnesium sulfate, the solvent was removed under reduced pressure to give crude (2-isocyanoethyl)benzene as a viscous, dark brown oil. The crude product was sealed and stored in the freezer overnight.

Reaction Setup:
Start of reflux
Reaction mixture at 1.5 hours
Reaction mixture at 2.5 hours
Reaction mixture at 4 hours.
Emulsion prior to dilution
Crude product as an oil


Crude 250 mL round bottom flask weight: 100.803 g.

02/12/11

Purification
The crude oil was loaded on a 5 cm silica gel column (diameter 3 cm) and eluted with neat EtOAc to form a yellow-brown oil. The product seemed too dark in colour and the yield was higher than expected. The product was passed through another silica gel column (30 cm, 3 cm diameter, neat EtOAc). Clearer separation was achieved and the yellow filtrate was collected. The solvent was removed under reduced pressure to give KAB4-1 as a yellow-brown oil (16.6 g, 66 %).

Second column
Dilute purified KAB4-1


Flask + Product: 117.365 g.
Yield: 16.6 g
Percentage yield: 66 %

NMR Data
H-NMR of KAB4-1 (CDCl3)

(UPLOAD RAW HNMR DATA FOR KAB4-1)

***Procedure finished. The method for the isonitrile preparation was straightforward and agreed with the literature procedure outline and expected yield.

References
[1] H. Cao, H. Liu, A. Doemling, Chemistry-a European Journal 2010, 16, 12296. DOI: 10.1002/chem.201002046


Forward Experiments
Preparation of the PZQ analogue Ugi-intermediate (KAB5-1)
Preparation of the N-benzoyl PZQ analogue Ugi-intermediate (KAB6-1)

Related Experiments
Preparation of (2-isocyanoethyl)benzene (MNR21-1)

Links
Informal commentary
Linked Posts
Attached Files
24th November 2011 @ 01:08
Further attempts to sort out the problems of the Newman-Kwart Rearrangement

Continuing on from MNR20-1 - MNR20-6

MNR20-1%20scheme.png

MNR20-1%20table.PNG

Hazard and Risk Assessment:
As for HIRAC-MNR17

Procedure
MNR20-7

MNR19-1 (70 mg, 0.15 mmol) in a sealed tube round was placed in a pre heated sand bath at 240-250°C (Heidolph hot plate). After 1 hour the tube was removed from the heat and allowed to reach room temperature.

Crude 1H NMR showed conversion but not complete or clean.

**NMR is named 20-6 but is actually 20-7**

25-75% EtOAC/Hex.

Product came out as very weak spots in fracs 15-23 - 15 mgs

1H NMR mnr20-7 frac 15-23

repeat above conditions with slightly longer reaction time.

Column, 5/12/11

Lab book page complete, MNR

Procedure
MNR20-8

MNR19-1 (40 mg, 0.09 mmol) in a sealed tube round was placed in a pre heated sand bath at 240-250°C (Heidolph hot plate). After 2 hours the tube was removed from the heat and allowed to reach room temperature.

Column:- 25-75% EtoAC/Hex

Frac 10-23
frac10-23

Frac 41-57 - mass?
product 12H peak resolves better at 500MHz

Lab book page complete, MNR
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24th November 2011 @ 01:04
Slight confusion over the labelling of sample vials. Both MW36 and MW29 are compound 1a (Scheme 1).

Scheme 1
Outline_Pictet_Spengler.png

Procedure
Scheme%203-1.png

Similar reaction conditions for the preparation of MNR11-9 were applied to MW29-4.

Related Experiments
Acid-mediated Pictet-Spengler of MW29 to give rac-PZQ MNR13-1
Attempts to the acid-mediated Pictet-Spengler cyclization of the ‘Ugi intermediate’ MW29 (MW31)

Hazard and Risk Assessment
HIRAS 3-1



Preparation of KAB3-1:
KAB3-1%20table.tjpg
MW29-4 (42 mg) was dissolved in toluene (0.6 mL) before the addition of methanesulfonic acid (0.001 mL) to give a clear, colourless solution. The reaction mixture was heated in an oil bath, 116C at 3:00 pm. The temperature of the oil bath fluctuated between 113-120C until the reaction mixture was removed from heat at 4:00 pm - total 1 hour at reflux. The solution had turned yellow accompanied by a dark red, non-miscible substance.

A sample of KAB3-1 was extracted from the reaction mixture then quenched with saturated sodium bicarbonate. TLC was performed on the KAB3-1 extraction against the starting material MW29-4, in ethylacetate:hexane, 3:1 with a KMnO4 stain (Photo 2).

KAB3-1 1st TLC 75%

Photo 2. From top to bottom: (1)MW29-4, (2)MW29-4 and KAB3-1, (3)KAB3-1


It was difficult to see what was product, starting material or byproduct. The TLC was repeated (ethylacetate:hexane, 3:1; KMnO4 stain) with the addition of racemic praziquantel (PZQ) (Photo 3).

KAB3-1 TLC 75 %

Photo 3. From left to right: (1)MW29-4, (2)MW29-4, KAB3-1 and PZQ, (3)KAB3-1, (4)PZQ

It was still difficult to determine the progress of the reaction based on this TLC. The TLC was repeated, reducing polarity of the mobile phase, to promote separation of any close eluting spots. Another TLC was performed in ethylacetate:hexane, 1:1 then stained with KMnO4 (Photo 4).

50% TLC

Photo 4. From left to right: (1)MW29-4, (2)MW29-4, KAB3-1 and PZQ, (3)KAB3-1, (4)PZQ

The TLC in 50% ethylacetate indicated consumption of the MW29-4 starting material and suggested the formation of the PZQ. The second fastest spot in KAB3-1 was unknown.

The KAB3-1 reaction mixture was quenched with saturated sodium bicarbonate (10 mL) then extracted with ethylacetate (3 x 15 mL) (Photos 5-7). The organic layers were combined then dried over magnesium sufate to give a clear yellow solution. The solvent was removed under reduced pressure to give crude KAB3-1 (PZQ) as a dark brown-red oil (28.6 mg)

Quenching KAB3-1

Photo 5: Quenching of KAB3-1 with sodium bicarbonate before the first extraction. The non-miscible red layer was clearly visible at the organic-aqueous interface.
KAB3-1 2nd Extraction

Photo 6: The aqueous layer of KAB3-1 just before the second extraction with ethylacetate.
Final KAB3-1 extraction

Photo 7: The less coloured organic layer above the orange-brown aqueous layer, just after the final extraction

Empty flask: 33.4408 g
Flask + sample: 33.4694 g

CRUDE YIELD of KAB3-1 (PZQ): 28.6 mg

25/11/11
Attempt at purification by flash column chromatography resulted in poor separation (Ethylacetate in hexane 20-100%). Monitoring separation by TLC was difficult due to high dilution of KAB3-1.
The column was flushed with neat methanol and all fractions combined. The solvent was removed to give crude KAB3-1 (~30 mg). Little to no product was lost.

28/11/11 - Second attempt at purification of KAB3-1
The recovered KAB3-1 was purified by flash column chromatography (silica gel 6 cm, diameter 1 cm in ethylacetate:hexane, 1:1).
Fourteen fractions were collected. Thin Layer Chromatography (in ethylacetate:hexane, 1:1) indicated elution of product in fractions 1-3 and 5-8. Fractions 1-3 and 5-8 were combined to give KAB3-1a and KAB3-1b, respectively. Both solutions were concentrated under reduced pressure.

Concentrated KAB3-1b was a thick yellow oil. TLC of KAB3-1b was performed against racemic PZQ and both had Rf of ~0.4.
KAB3-1b was put under high vacuum for 3 hours before storage in the freezer overnight.

KAB3-1a was concentrated under reduced pressure to give a yellow oil.

NMR Data
KAB3-1a NMR



29/11/11

KAB3-1b was removed from the freezer - no crystallisation. The solvent was removed from KAB3-1a under reduced pressure.

YIELD
Fractions 1-3 KAB3-1a: 10 mg
Fractions 5-8 KAB3-1b: 3 mg

TLC fractions

Repeat TLC of KAB3-1a and KAB3-1b. From left to right: (1) KAB3-1b, (2)KAB3-1b duplicate, (3)racemic PZQ, (4)KAB3-1b, rac-PZQ and KAB3-1a, (5)KAB3-1a

The solvent was removed from KAB3-1a under reduced pressure and the sample placed under high vacuum. NMR analysis of KAB3-1a and KAB3-1b pending.


Preparation of KAB3-2

24/11/11

The reaction was prepared as above, MW29-4 (24 mg), toluene (0.6 mL) and methanesulfonic acid (0.001 mL). The clear, colourless mixture was stirred at room temperature from 4:15 pm (Photo 8).

KAB3-2 at time zero

Photo 8. KAB3-2 at time zero

At 1.5 hours after the reaction start, a sample of KAB3-2 was extracted and quenched in saturated sodium carbonate before TLC analysis in ethylacetate:hexane, 1:1. The TLC showed no spots corresponding to the starting material (MW29-4) or the expected product KAB3-X (PZQ). The large, fast eluting smear present in the KAB3-1 and all MW29-4 TLCs was present in the KAB3-2 TLC. The reaction mixture was left to stir at room temperature overnight.
Unlike KAB3-1, the reaction mixture of KAB3-2 at 1.5 remained mostly clear and colourless. However, there was some formation of a brown precipitate.

KAB3-2 after 1.5 hours

Photo 9: KAB3-2 after 1.5 hours.

29/11/11

The reaction mixture removed from heat, diluted with EtOAc (5 mL) then quenched with saturated sodium bicarbonate (6 mL). Following separation of the organic fraction, the aqueous layer was extracted with EtOAc (2 x 5 mL). The combined organic fractions were dried over magnesium sulfate, before removal of the solvent under reduced pressure. Crude yield: 13.1 mg.


***Procedure finished. Difficult to make any conclusions from this experiment due to the low mass of product obtained (I'm still getting used to working with tiny amounts). However, procedure was utilised in future experiment KAB3-5 in attempt to push the peptide acetal Ugi intermediate to PZQ product.

Following experiments
The reflux conditions were adapted in:
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)
Acid-mediated Pictet-Spengler reaction to give PZQ (KAB3-4) and the N-benzoyl PZQ analogue (KAB7-2)
Acid-mediated Pictet-Spengler reaction to give PZQ (KAB3-5)


No further characterisation of the product has been performed as of 08/12/11 due to low quantity of product. The NMR recorded of the samples were too dilute. It is unknown what the outcome of the experiment was.
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23rd November 2011 @ 23:22
**I've been locked out of the previous post due to mix up in google accounts**

Acid-mediated Pictet-Spengler of MW51 to give N-benzoyl-derivative of PZQ MNR14-1 - comparing the rates of reaction versus the activated examples MNR11, MNR12 and MNR13-1

MNR14-1%20scheme.png

MNR14-1%20table.PNG

Hazard and Risk Assessment:
As for MNR12-2

Procedure
MW51 (95 mg, 0.28 mmol) was stirred with methanesulfonic acid (1.82 mL, 28 mmol) at room temperature and after 1 hour (first TLC) all the starting material had been consumed. The reaction was quenched with saturated sodium carbonate and extracted with EtOAc (20 mL x2). The organic fractions were dried over magnesium sulfate, filtered and concentrated under reduced pressure.

Crude - 60 mg as a yellow oil that showed signs of slowly crystallizing after 24 hours

1H MNR14-1 Crude
1H MNR14-1 vs MNR12-1
Linked Entries
Attached Files
22nd November 2011 @ 07:30
Samples MNR12-1 and MNR12-2 were combined and purified by column chromatography (silica gel, ethyl acetate:hexane 50-100 %). The products KAB2-1a and KAB2-1b were observed by TLC in ethylacetate before the fractions were combined and the solvent removed under reduced pressure.

Scheme%202-1.png

Yield
KAB2-1a: 41 mg (fractions 77-88)
KAB2-1b: 58 mg (fractions 7-12)
Everything else: 95 mg

Total recovery: 194 mg
Amount loaded: 180 mg

TLC of crude MNR12-1 and MNR12-2 in ethyl acetate
Stain: KMnO4
Crude TLC

TLC of fractions 77-89



Characterisation

22/11/11: H-NMR was recorded for KAB2-1a. Very messy - Needs further tweaking/processing, possibly changing temp and diluting further. Also has ethyl acetate contamination.
HNMR KAB2-1a

note: The spectrum similar to the SciFinder predicted H-NMR.

23/11/11: The NMR sample prepared yesterday was further diluted in CDCl3 and the spectrum re-recorded. It was slightly cleaner. Need to process the data, results pending.
The KAB2-1 samples in CDCl3 were recombined and the solvent removed under reduced pressure. giving a clear oil, which was placed in the freezer overnight to promote crystallisation.

24/11/11:
8:17 am - No crystallisation yet
Photo.jpg

The KAB2-1 sample was removed from the freezer and put under high vacuum. The remaining fractions (1-6, 13-76 and 89-111) were combined and the solvent removed to give a slightly yellow solid with a mass of 95 mg. The total recovery was 108 % of the amount loaded. Possible silica contamination from the column.

7:11 pm - Sample KAB2-1a was returned to freezer


24/11/11:
8:17 am - KAB2-1a Still no crystallisation.

28/11/11:
8:13 am - KAB2-1a Still no crystallisation. Sample removed from freezer and left at room temperature.

28/11/11:
Sample no longer a gel, but a yellow solid. Attempt at recrystallisation from toluene and hexane.

08/12/11:
Formation of small yellow crystals. Flask kept at room temperature.

15/12/11:
Crystallisation complete. The small yellow crystals were washed in hexane then filtered.

16/12/11:
1H-NMR recorded.

03/01/12:
Yield: 27.8 mg, 19.3%
Rough m.p.: 109-120 C
Final KAB2-1


Recovery and yield for the purification procedure was poor. As of 23/01/12, no characterisation performed on the recovered crystals.

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Attached Files