Author Archives: Brett Pogostin

Synthesis of 1-(6-chloropyrazin-2-yl)-2,2,2-trifluoroethan-1-one (BPVR 1-1) Attempt 2

O=C(C1=NC(Cl)=CN=C1)C(F)(F)F

 

Reference

Holsinger, L.J., Compounds that Inhibit Protease Cathepsins and HCV Replication. U.S. Patent 2008080785, April 30, 2009.

 UNSUCCESSFUL
 

Substance

Amount

Mol. wt.

mmols

Equiv.

Starting Material

1 g

172.57

5.79

1

trimethylsilyltrifluoro-methane

0.70 g

142.2

4.92

0.85

caesium fluoride

0.880

151.9

5.79

1

tetra-butylammoniun fluoride (1M in THF)

7.25 mL

261.46

7.25

1.25

Concentrated Acetic Acid (17.4 M)

2.8 mL

 

49.2

8.5

Product

 

1-(6-chloropyrazin-2-yl)-2,2,2-trifluoroethan-1-one

1.033 g (Theoretical)

 

0.1127 mg

(Actual)

209.98

4.92 (Theoretical)

 

0.54

(Actual)

0.85 (Theoretical)

 

11% Yield

(Actual)

 

Procedure:

(11/30/16)

 
  1. Dissolved starting material in dimethoxy ethane (15 mL).

  2. Dried CsF (0.88g) under vacuum while heating with a heat gun.

  3. Add trimethylsilyltrifluoro-methane (0.82 g) and CsF (0.880 g) to the starting material solution.

  4. Stirred at 4 degrees celsius for 36 hours.
    (12/2/16)

  5. A TLC was taken to see if the reaction was complete.

  6. The reaction was then concentrated at room temperature in high vac to a brown oil.



    (12/7/16)

  7. The concentrated product was then dissolved in 10 mL of THF, 2.8 mL acetic acid, and treated with 7.25 mL of tetrabutylammonium fluoride.

  8. Reaction was aged for 3 hours and then stopped by diluting the reaction with ethyl acetate (20 mL). A TLC was taken after 3 hours but no change was observed between the starting material before the workup and the product after the workup.

  9. The product was then gravity filtered to remove excess CsF.

  10. Product was washed three times with dilute aqueous sodium bicarbonate solution (0.25M, 100 mL portions).

  11. Washed with brine (100 mL).

  12. Dried over anhydrous magnesium sulfate, then filtered.

  13. Concentrated in vacuo.

1;1 Ethyl Acetate : Hexanes; S → Starting Material, C→ Co-Spot, P → Product
(12/8/16)

  1. Triturated brown oil product in a minimal amount of dichloromethane and hexanes to yield a brown solid.

    1. Yield: 0.1127 g (11%)

 

Analysis: In order to determine the identity of the product, GCMS and NMR were run.

 

GCMS:

 
[Untitled].pdf

The Peak at 207 m/z is the same that was observed last attempt. The mass of the expected product is 209, thus this data suggests that the synthesis was not successful.

 

NMR:

 

Both Carbon and PRoton NMR do not match any the expected spectra for the desired product, which further suggests that the synthesis was not successful.

 

Conclusions:

 

Decreasing the temperature of the reaction and decreasing the molar equivalents of the reactants did not improve the outcome of the reaction. Looking back at the previous group that worked on this project, their data suggests that they too were not successful in making this compound. We would suggest that future attempts to synthesize “The Burns” take another synthesis route. One possible synthesis is proposed below:

 

other thing.jpg

 

The reference for this reaction didn’t perform this reaction exactly; however, it might still work. Another option is to come up with a complete new total synthesis for “The Burns” compound.

 

Deprotection attempt 2 of tert-butyl 3-(5-(2-phenyl-2-((tetrahydro-2H-pyran-2-yl)oxy)ethoxy)-[1,2,4]triazolo[4,3-a]pyrazin-3-yl)piperidine-1-carboxylate (MRLS 9-1)


deprotect.jpg

Compound

Mw (g/mol)

mmol

mL

g

equiv

Starting Material

526.6501

0.09494

 

50mg

1

HCl-Ether

38

0.0312

0.156 (2M HCl)

 

3.3

Product

Theoretical Values

339.17

0.09494

 

32.2mg

1

dioxane

88.11

 

2 mL (4M)

 

3.3

Product Actual Value

339.17

0.07371

 

25.0 mg

78% Yield

 

November 30, 2016

Procedure:

  1. Dissolve 50 mg of the starting material in 500 uL of anhydrous diethyl ether and place system under argon.

  2. Add 2M HCl-Ether (0.1 mL) dropwise to the solution while stirring.

  3. Age solution for 2 hours at room temperature and the reaction was monitored with TLC.

  4. Left in fridge overnight.

the nemesis deprot round 2.jpg

100% EtOAc solvent system, silica gel plate, UV visualization

S → Starting Material; C → Co-spot; P → Product.

This TLC is similar to the first deprotection we did, which we deemed unsuccessful in taking out that BOC group, though there was success in knocking out the THP. This can be determined by comparing the S and C to the P. The S and C show a dark spot higher up on the plate; P shows one lower with a faint smear above it. This suggests that the THP group was successfully knocked out and the BOC group only partially (this agrees with our NMR and GCMS results from the first deprotection). Another TLC was done to confirm this hypothesis and determine if the reaction should be hit harder to take out both the THP and BOC group.

  the nemesisdeproround2.jpg

100% EtOAc solvent system, silica gel plate, dipped in ninhydrin and treated with heat

S1→ Original Purified Starting Material; S2→ Starting Material from First Deprotection; C → Co-spot; P → Product.

Treating the TLC plate with ninhydrin and heat results in two dark spots on the base line of S2, and C and a faint brown spot on the base line of P. The difference in intensity can be attributed to concentration. The presence of these spots after being treated with heat suggest that the BOC group was successfully knocked off, and resulted in the formation of an amine, which is the desired product. The BOC was only successful knocked off with heat on the plate. To do this on a larger scale, these results suggests the deprotection reaction must be hit harder; the reaction must be done under heat with the addition of dioxane.

 

December 5, 2016

We were unsuccessful in removing the BOC with extra time . This suggests the reaction needs to be hit harder, specifically with the addition of dioxane and ran in an oil bath.

  1. Added 2 ml of 4M Dioxane and 0.1 mL of HCl  dropwise to the product while stirring and place in an oil bath at 50 degrees Celsius. System was placed under argon.  The reaction was monitored with TLC.

  2. After 2 hours at 50 degrees, TLC indicated that there was still starting material. The temperature was raised to 102 degrees to induce reflux. The reaction was then left to reflux for 45 minutes.

  3. After 45 minutes the reaction turned brown and a TLC was taken. The results of the TLC and a ninhydrin test suggest that the complete deprotection was successful due to the disappearance of the high Rf spot observed in previous attempts.

 

December 7, 2016

Mass Spec:

 
Deprotection.pdf
 

The peak at m/z 340 suggests that we successfully removed the Boc protecting group that we had trouble removing in the past. It is possible, however, some residual Boc protecting groups were removed in the mass spec instrument, so the product was also analyzed with proton and carbon NMR.

 
December 8, 2016 

NMR:

Still to be determined

 

Synthesis of 1-(6-chloropyrazin-2-yl)-2,2,2-trifluoroethan-1-one (BPVR 1-1)

O=C(C1=NC(Cl)=CN=C1)C(F)(F)F

 

Reference

Holsinger, L.J., Compounds that Inhibit Protease Cathepsins and HCV Replication. U.S. Patent 2008080785, April 30, 2009.

 
 

Substance

Amount

Mol. wt.

mmols

Equiv.

Starting Material

1 g

172.57

5.79

1

trimethylsilyltrifluoro-methane

1 g

142.2

7.032

1.35

caesium fluoride

0.880

151.9

5.79

1

tertbutylammoniun fluoride

1.514 g

261.46

5.79

1

Product

 

1-(6-chloropyrazin-2-yl)-2,2,2-trifluoroethan-1-one

1.216 g (Theoretical)

 
 

(Actual)

209.98

5.90 (Theoretical)

 
 

(Actual)

1 (Theoretical)

 
 

(Actual)

 

Procedure:

(11/16/16)

  1. Dissolved starting material in dimethoxy ethane (15 mL).

  2. Dried CsF (1.435g) under vacuum while heating with a heat gun.

  3. Add trimethylsilyltrifluoro-methane (1 g) and CsF (0.880 g) to the starting material solution.

  4. Stirred at room temperature overnight.
    (11/17/16)

  5. The reaction was then concentrated under reduced pressure at room temperature.
    (11/19/16)

  6. The concentrated product was then dissolved in 7.2 mL of THF, 3 mL acetic acid, and treated with 1.514 g of tetrabutylammonium fluoride.

  7. Reaction was aged for 3 hours, while progress was monitored with TLC, and then stopped by diluting the reaction with ethyl acetate (20 mL).

  8. Washed three times with dilute aqueous sodium bicarbonate solution (0.25M, 100 mL portions).

  9. Washed with brine (100 mL).

  10. Dried over anhydrous magnesium sulfate, then filtered.

  11. Concentrated in vacuo.
    (11/21/16)

  12. Attempted to triturated in dichloromethane and hexanes, but it was not successful. TLC was taken showing that product contained significant impurities.
    IMAG0104.jpg
    50% hexanes in EtOAc solvent system, silica gel plate, UV visualization.
    Bottom is starting material, middle is a co-spot and the top is the product.

    (11/21/16)

  13. Attempted to separate the different spots observed in the TLC on the biotage. Several fractions collected containing mixtures of different products.

  14. All the fractions were spotted with TLC and those containing only one product were separated. Fraction 20 which may contain the starting material was separated and concentrated in vacuo. Fractions 23, 24, and 25, which may contain the product, were concentrated. And fractions 12-16 were also concentrated which contain an unknown byproduct. Fractions 21 and 22, which contained a mixture of two products were concentrated together for further separation.
    Notes: Some other oil seemed to have made its way into the fraction 20 concentrate, so it may need further purification.


 

(11/28/16)

  1. Prepared fraction 23-25 and fraction 20 for NMR. Both samples were readily dissolved in chloroform.

  2. The fractions were dissolved in acetonitrile and analyzed via Gas Chromatography.

    Fractions 23 - 25:


    Fraction 20:

 
 
  1. Fractions 23-25 and 20 were dissolved in chloroform and analyzed via NMR

Fraction 23-25  Carbon NMR

VRBPBurns2325CarbonNMRchloroform.JPG

Fraction 23-25 Proton NMR

VRBPBurns2325ProtonNMRchloroform.JPG

 
 
 
 
 

Fraction 20 Carbon NMR

VRBPBurns20CarbonNMRchloroform.JPG

Fraction 20 Proton NMR

VRBPBurns20ProtonNMRchloroform.JPG

 
 

Though it was predicted  Fraction 23-25 would be the location of our main product, NMR results suggest product is actually present in Fraction 20. There are many unidentifiable peaks present in both NMR spectra of Fraction 23-25 compared to those of Fraction 20. Proton NMR of Fraction 20 shows all anticipated peaks, though the strong peak at 4 ppm remains to be identified. Carbon NMR also came out as expected for desired product except for small unidentifiable peaks at approximately 50 ppm and 165 ppm. These impurities suggests that a synthesis of 1-(6-chloropyrazin-2-yl)-2,2,2-trifluoroethane-1-one should be attempted again, at a lower temperature and lower equivalence of trimethylsilyltrifluoro-methane.

Deprotection of tert-butyl 3-(5-(2-phenyl-2-((tetrahydro-2H-pyran-2-yl)oxy)ethoxy)-[1,2,4]triazolo[4,3-a]pyrazin-3-yl)piperidine-1-carboxylate


deprotect.jpg
 
 
 

Compound

Mw (g/mol)

mmol

mL

g

equiv

Starting Material

526.6501

0.09494

 

50mg

1

HCl

38

0.0312

0.156 (2M HCl)

 

3.3

Product

Theoretical Values

339.17

0.09494

 

32.2

1

 

November 14, 2016

Procedure:

  1. Dissolve 50 mg of the starting material in 500 uL of anhydrous diethyl ether and place system under argon.

  2. Add 2M HCl-Ether (0.1 mL) dropwise to the solution while stirring.

  3. Age solution for 30 mins and the reaction was monitored with TLC.

IMAG0102.jpg

 

100% EtOAc solvent system, silica gel plate, UV visualization

S → Starting Material; C → Co-spot; P → Product

  1. The TLC of the reaction indicated that it was not complete, so another 500 uL of anhydrous ether and 50 uL of 2M HCl-ether was added to the reaction.

  2. The solution was then aged for another 30 minutes.

  3. TLC of the reaction was taken after the additional 30 minutes. Starting material was present, indicating the  reaction incomplete (presence of a salt). 500 uL of anhydrous ether was added to the reaction to ensure it remained in solution.

100% EtOAc solvent system, silica gel plate, UV visualization

S → Starting Material; C → Co-spot; P → Product

  1. The reaction was then gravity filtered and washed with diethyl ether to isolate the product. The product was a fine yellow solid.

 

November 16, 2016

Procedure

  1. The fine pale yellow solid was then mixed with diethyl ether (~8 mL) in a small r.b. Flask.

  2. Because the product was found to be insoluble in diethyl ether, it was put in a sonicator bath to speed up dissolution/resuspension.

  3. The mixture was left to settle in the r.b. A pale yellow, very fine solid settled  at the bottom of the flask.

  4. Vacuum filtration was done to separate the solid product from the diethyl ether. It was then washed once with diethyl ether to yield  0.0125g of final product.

  5. In preparation for NMR, 1 mL of methanol was added to 5 mg of the final product. (The final product (5 mg) was found to be insoluble in (1 mL) D2O, even after being put in the sonicator; this resulted in resuspension rather than dissolution).

  6. NMR was run to determine presence of a salt and the identity of the product.

Purification and Analysis of tert-butyl 3-(5-(2-phenyl-2-((tetrahydro-2H-pyran-2-yl)oxy)ethoxy)-[1,2,4]triazolo[4,3-a]pyrazin-3-yl)piperidine-1-carboxylate (MRLS 9-1)

11/9/16

Purification:

 
  1. The 550 mg of MSLR 9-1 was dry loaded onto silica and the product was purified using the biotage using a solvent system of 100% ETOAc.

  2. Fractions containing product were checked using TLC. TLC shows that the product was successfully isolated. In 3 fractions, high Rf impurities might be present in low concentration, however it is unclear and this will be addressed if the NMR looks messy.

  3. The product was concentrated in vacuo. Yield: 0.145 grams

100% EtOAc solvent system, silica gel plate, UV visualizationTLC100ethylacetatemarcolincoln9-1.jpg

The numbers at the bottom of the TLC plates are the factions we identified our product to be in.  The curve seen between the factions on the TLC suggests there are diastereomers present in our final product.

 
 

11/10/2016

4. Used HNMR to determine if THP and BOC groups were present in the product.

The tall peak at approximately 1.5 ppm suggests the presence of a BOC group (the 9H on the quaternary carbon at the end). The presence of peaks between 3-4 pm, as well as others around 1 ppm suggest THP is present .

 Conclusions:
 
It appears that the protecting groups are present. A deprotection step will be done to see if the final product is successfully synthesized.

10/31/16-11/3/16 Synthesis of 1-(6-chloropyrazin-2-yl)-2,2,2-trifluoroethan-1-one (EVKCN 1-3)

Purpose: In order to work out the issues seen in the reductive amination step (see Lab Notebook), more precursor needs to be prepared.

10-31-16 step 1 reaction.JPG

Image courtesy of Eliana von  Krusenstiern and Claudia Nguyen. Values on table represent both what we did and the yeild values that Claudia and Eliana produced when they ran this reaction.

Procedure:

(10/31)

  1. Malononitrile (0.887 g, 13.42 mmol) was dissolved in 85 mL of dry THF under argon.

  2. NaH in mineral oil suspension (0.45g, 60% NaH by mass) was added slowly to the mixture prepared above and the solution was allowed to age for 30 mins.

  3. 2,6-dichloropyrazine (1g, 6.71mmol) was dissolved in dry THF (25 mL). Some white solid impurity remained undissolved, which was also reported by Eliana and Claudia in their lab notebook. The solution was added dropwise to the aged solution under argon.

  4. (11/2/16)
    The mixture was green-yellow and was placed in Reflux at 60 ℃ for 5 hours. The solution dried out and another 25 mL of dry THF was added and the solution refluxed for another 5 hours. The reactions was losing solvent to a hole in the glassware.

  5. (11/3/16)
    To acidify the solution, 20 mL of 1M HCL was added to the mixture. The solution turned deep red.

  6. Ethyl acetate (50 mL) was added to improve separation between the organic and aqueous layers, and the organic layer was separated from the aqueous layer.

10/31/16-11/7/16 Synthesis of 1-(6-chloropyrazin-2-yl)-2,2,2-trifluoroethan-1-one (EVKCN 1-3)

Purpose: In order to work out the issues seen in the reductive amination step (see Lab Notebook), more precursor needs to be prepared.

10-31-16 step 1 reaction.JPG

Image courtesy of Eliana von  Krusenstiern and Claudia Nguyen. Values on table represent both what we did and the yeild values that Claudia and Eliana produced when they ran this reaction.

Procedure:

(10/31)

  1. Malononitrile (0.887 g, 13.42 mmol) was dissolved in 85 mL of dry THF under argon.

  2. NaH in mineral oil suspension (0.45g, 60% NaH by mass) was added slowly to the mixture prepared above and the solution was allowed to age for 30 mins.

  3. 2,6-dichloropyrazine (1g, 6.71mmol) was dissolved in dry THF (25 mL). Some white solid impurity remained undissolved, which was also reported by Eliana and Claudia in their lab notebook. The solution was added dropwise to the aged solution under argon.

  4. (11/2/16)
    The mixture was green-yellow and was placed in Reflux at 60 ℃ for 5 hours. The solution dried out and another 25 mL of dry THF was added and the solution refluxed for another 5 hours. The reactions was losing solvent to a hole in the glassware.

  5. (11/3/16)
    To acidify the solution, 20 mL of 1M HCL was added to the mixture. The solution turned deep red.

  6. Ethyl acetate (50 mL) was added to improve separation between the organic and aqueous layers, and the organic layer was separated from the aqueous layer.

  7. (11/4/16)
    The organic phase was separated and volatiles removed in vacuo.

  8. The residue was partitioned between H2O (150 mL) and Et2O (150 mL) to wash away inorganic salts and excess malononitrile. The organic layer was then washed with H2O and brine and then dried. The solution was a deep red/orange color.

  9. Significant product was lost during the extraction due to accidental disposal. A brown solid contaminant was observed in the organic layer.

  10. (11/7/16)
    The brown solid observed above was removed via vacuum filtration and the solution was concentrated in vacuo. Our final product was a dark brown liquid. Water vapor around the neck of the r.b. was seen in vacuo, which indicates the presence of some water contamination in the final product (0.13g).

Observation and Analysis:

TLC:

After Reflux:

10% methanol in EtOAc solvent system, silica gel plate, UV visualization

Left = After 10 horus reflux, Right = After 5 hours reflux

Rf(SM) = 0.75, Rf(Prod)= 0.3

The TLC suggests that the reflex reaction was successful and product was formed.

After Extractions:


Conclusions:


The sysnthesis was unsuccessful. TLC analysis shows that whatever was isolated is neither our starting material or our product. The product was likely lost due to error during the extraction step as noted above.

10/31/16-11/7/16 Synthesis of 1-(6-chloropyrazin-2-yl)-2,2,2-trifluoroethan-1-one

Purpose: In order to work out the issues seen in the reductive amination step (see Lab Notebook), more precursor needs to be prepared.

10-31-16 step 1 reaction.JPG

Image courtesy of Eliana von  Krusenstiern and Claudia Nguyen. Values on table represent both what we did and the yeild values that Claudia and Eliana produced when they ran this reaction.

Procedure:

(10/31)

  1. Malononitrile (0.887 g, 13.42 mmol) was dissolved in 85 mL of dry THF under argon.

  2. NaH in mineral oil suspension (0.45g, 60% NaH by mass) was added slowly to the mixture prepared above and the solution was allowed to age for 30 mins.

  3. 2,6-dichloropyrazine (1g, 6.71mmol) was dissolved in dry THF (25 mL). Some white solid impurity remained undissolved, which was also reported by Eliana and Claudia in their lab notebook. The solution was added dropwise to the aged solution under argon.

  4. (11/2/16)
    The mixture was green-yellow and was placed in Reflux at 60 ℃ for 5 hours. The solution dried out and another 25 mL of dry THF was added and the solution refluxed for another 5 hours. The reactions was losing solvent to a hole in the glassware.

  5. (11/3/16)
    To acidify the solution, 20 mL of 1M HCL was added to the mixture. The solution turned deep red.

  6. Ethyl acetate (50 mL) was added to improve separation between the organic and aqueous layers, and the organic layer was separated from the aqueous layer.

  7. (11/4/16)
    The organic phase was separated and volatiles removed in vacuo.

  8. The residue was partitioned between H2O (150 mL) and Et2O (150 mL) to wash away inorganic salts and excess malononitrile. The organic layer was then washed with H2O and brine and then dried. The solution was a deep red/orange color.

  9. Significant product was lost during the extraction due to accidental disposal. A brown solid contaminant was observed in the organic layer.

  10. (11/7/16)
    The brown solid observed above was removed via vacuum filtration and the solution was concentrated in vacuo. Our final product was a dark brown liquid. Water vapor around the neck of the r.b. was seen in vacuo, which indicates the presence of some water contamination in the final product (0.13g).

Observation and Analysis:

TLC:

After Reflux:

10% methanol in EtOAc solvent system, silica gel plate, UV visualization

Left = After 10 horus reflux, Right = After 5 hours reflux

Rf(SM) = 0.75, Rf(Prod)= 0.3

The TLC suggests that the reflex reaction was successful and product was formed.

After Extractions:

20161107_150253.jpg

10% methanol in EtOAc solvent system, silica gel plate, UV visualization

Conclusions:


The sysnthesis was unsuccessful. TLC analysis shows that whatever was isolated is neither our starting material or our product. The product was likely lost due to error during the extraction step as noted above. We found a commercially available source to purchase a further intermediate, so we will continue with that.