6.17: Glovebox and Impurity Sensors

1. Bringing Items into the Glovebox

1. Make sure that the item(s) to be brought in have been oven-dried (if glassware), and that containers are open.
2. Check the antechamber log to ensure that it is empty.
3. Fill the antechamber, either manually or electronically. Once filled with 1 atm of inert gas, close the inlet valve to isolate the chamber.
4. Open the antechamber to the outside, and place the items in the chamber.
5. Close the chamber, and evacuate (manually or electronically).
6. Fill in the log. Typically, users include their initials, items, and times of each cycle.
7. After the pressure dial reaches the minimum pressure, leave the antechamber under dynamic vacuum for 5 min for a small antechamber, and 20 min for a large antechamber.
8. Refill the antechamber with inert gas; typically, users will backfill to ~ 0.75 atm, as the inlet valve connects the main chamber to the antechamber.
9. Evacuate, and note the time.
10. Repeat steps 1.8-1.9, so that in total the antechamber has been evacuated 3x.
11. After 3 cycles, fill the antechamber with inert gas, and close off the insert gas supply.
12. Open the antechamber from inside the glovebox, and bring the items into the antechamber.
13. Close the antechamber door, and evacuate the chamber. When the glovebox is in its resting state, the chambers should be left under dynamic vacuum.
14. Note on the log that the procedure is complete, so that other users know that the antechamber is free.

2. Removing Items from the Glovebox

1. Look at the logbook to see the status of the antechamber. Be sure that it is not in use, and that the last operation was to bring an item into the antechamber. If the last operation was to bring an item out, quickly fill/evacuate the antechamber 3x with inert gas. This is to ensure no residual air is present (Equation 1) when opening the antechamber to the glovebox.
2. Fill the antechamber with inert gas, and close the valve connecting the inert gas supply to the chamber.
3. Open the antechamber from the inside of the glovebox.
4. Load the items into the chamber, and close the door.
5. From the outside of the glovebox, open the antechamber door and remove the items.
6. Evacuate the chamber.
7. Note that items were removed and the time on the logbook.

3. Ensuring a Good Working Environment

1. Testing the environment
   1. Turn off the circulator.
   2. Turn off any fans in the main glovebox chamber.
   3. Open a bottle of diethylzinc solution in hexanes (frequently 1.0 M).
   4. Gently swirl the bottle to replace the gas atmosphere in the bottle with the atmosphere in the box. If smoke emerges from the bottle, this is an indication that O₂, water, or an ether solvent is present in the atmosphere. If the atmosphere is compromised, the source of unwanted impurities should be identified.
   5. Turn the purge on for 5 min.
   6. Turn the purge off and the circulator on.
2. Making the radical indicator
   1. Turn off the circulator.
   2. In the glovebox, weigh out 5 mg of benzophenone and transfer this to a 20-mL scintillation vial.
   3. Weigh out ~ 500-1,000 mg of sodium and transfer this to the scintillation vial. Cap the vial.
   4. Add 20 mL of dry tetrahydrofuran (THF) and a stir bar. Cap the vial.
   5. Turn the purge on for at least 15 min before turning the circulator back on.
   6. Let the reaction stir for 48 h, or until the solution turns into a dark, inky purple solution. The solution should go from colorless to blue to purple, and there should be excess sodium at the bottom of the vial. This should give a solution with ~ 1.4 mM radical.
3. Testing solvent with the di-radical indicator
   Note: The newly synthesized radical can be used to test for O₂ and water impurities in solvents.
   1. If testing an ether solvent, turn the circulator off. Some groups require that the circulator is turned off before opening any chemical in the box.
   2. Add one drop of the radical solution to 10 mL of the test solvent. Solvents that can be tested using the radical include THF, diethyl ether, toluene, benzene, hexanes, and pentane. The radical will react with chlorinated solvents, pyridine, and other solvents that react with alkali metals.
   3. Observe the color of the solution over 1-2 min. A dry solvent will hold the color of the ketyl radical indefinitely. Realistically, the sample should hold the color for at least 1-2 min. Positive test colors are given in Table 1 below.
   4. Close all solvent bottles and turn the purge on for at least 15 min. Turn the circulator on again.

Sensitive substances like organolithium- or organometallic compounds can violently react when exposed to oxygen or water from air. Hence, an inert working environment is required, which can be achieved by using a glovebox.

The glovebox is an important device used in many laboratories, which allows handling and storage of air- and moisture sensitive compounds.

Furthermore, it can be used to measure sensitive substances and carry out reactions.

This video will illustrate how to operate the glovebox, and how to synthesize an indicator to test for oxygen and water within dry solvents.

In general, a glovebox is comprised of a metal box with polycarbonate windows fitted with butyl gloves allowing for manipulation inside the box. Chemicals and supplies are brought into the glovebox via the antechambers, while sensors and a control panel are used for monitoring and regulations.

Furthermore, the functionality of a glovebox can be extended by extra equipment, ranging from vacuum hook ups to freezers for chemical storage.

The glovebox atmosphere is achieved using inert gas such as nitrogen. The box is gas-tight and run at positive pressure, which is controlled by electronically regulating the gas flow into the system.

The inert atmosphere is circulated through a catalyst bed, which is located below the glovebox.

The catalyst is comprised of molecular sieves and copper, which are used to maintain a low level of oxygen and moisture. Copper reacts with oxygen present in the atmosphere, while molecular sieves absorb water. The catalyst has to be regenerated on a regular basis by heating it under a stream of hydrogen and nitrogen gas to assure its activity.

Besides moisture and oxygen, various solvents can contaminate the catalyst. To avoid this, the glovebox chamber is isolated, when working with incompatible chemicals.

Additionally, contamination can be introduced through the antechamber, which must undergo multiple evacuation and purging cycles to remove as much air as possible. The fraction of air remaining can be calculated using this equation.

The content of moisture and oxygen inside the box or any dry solvent can be tested using chemical sensors. Diethylzinc is used to test for contamination inside the box, while sodium benzophenone is used for solvents.

Now that you are familiar with the basics, let's take a look at how to operate the glovebox and test for oxygen and water.

Before you start familiarize yourself with the instrument. For an in detail instruction of glovebox usage watch our video in the laboratory safety collection. Assure that glassware to be brought in has been oven-dried, and empty containers are open.

Check the antechamber log to make sure it is empty. Then, fill the antechamber with inert gas to 1 atm, and close the inlet valve to isolate the chamber.

Once the chamber is purged, open it from the outside, and place the items inside the chamber. Close the chamber, and evacuate it.

Fill in the log including initials, items, and times of each cycle, while the chamber is evacuating. When minimum pressure is reached, leave the antechamber under dynamic vacuum between 5-20 min.

Then, using the inlet valve purge the antechamber again, wait until 1 atm?is reached, and evacuate again. Note the time and repeat the cycle. Lastly, refill the chamber with N2 and close off the inert gas supply, when the purging process is finished.

Now you are ready to open the antechamber from inside the glovebox to bring the items in. Close the antechamber door when finished, evacuate it, and fill out the log.?

Check the logbook for the last status of the antechamber and that it is not in use. Repeat the purging process if the antechamber was used to bring out items as the last operation. Then, close the valve connecting the inert gas supply, once antechamber is filled.

Open the door from inside, load the items into the chamber, and close the door. Then open the chamber from outside and remove the items. Evacuate the chamber and fill out the logbook.

Now that you are familiar with the proper usage of a glovebox, let's examine how impurity sensors can be used to test for oxygen and water in the glovebox atmosphere and various solvents.

To test the glovebox atmosphere for oxygen and water levels, first turn off the circulator. Then, open a bottle of diethylzinc solution in hexanes inside the glovebox.

Gently swirl the solution to replace the gas atmosphere with the glovebox atmosphere inside the bottle. Any emerging smoke and white residue indicates oxygen, water, or an ether solvent present in the atmosphere. Then, purge the glovebox for 5 min, turn off the purge, and turn the circulator back on when finished.

In addition to testing the glovebox atmosphere, indicators can be used to test various solvents for oxygen and water impurities. First, turn off the circulator. Then, open the bottle of the desired solvent and transfer 10 mL into a scintillation vial. Add one drop of the ketyl radical solution to test the solvent and observe the color over 1-2 min.

If the solvent is dry, it will hold the purple color of the ketyl-radical indefinitely. If the color changes to blue and then to colorless, then the solvent has impurities. To finish, close all solvent bottles, purge the glovebox and turn the circulator back on.

The glovebox is widely used to handle air- and moisture sensitive materials to carry out reactions, spectroscopic analysis, or to store compounds under air free conditions.

For example, the ketyl radical, which is used to test solvents for water and oxygen, is synthesized using a glovebox. To carry out the synthesis start with turning off the circulator. Weigh out 5 mg of benzophenone into a 20 mL scintillation vial. Then, weigh out 0.5-1 g of sodium and transfer it to the same scintillation vial together with a stir bar. Add 20 mL of dry THF and cap the vial.

Turn the circulator back on, after purging the glovebox for 15 min. Stir the reaction for 48 h or until the color changes from colorless to blue to purple. Once purple is reached, the ketyl radical is ready to use.

Besides chemical indicators, the glovebox can be used for the synthesis of air sensitive compounds, such as 1,2-azaborines.?

In this example N-H-B-ethyl-1,2-azaborine is synthesized starting from N-TBS-B-Cl-1,2-azaborine using a glovebox and a Schlenk line. The isolated compound is then used to prepare a protein-ligand crystal complex with purified lysozyme mutants, and the protein-binding interactions are studied using X-ray diffraction analysis.

You've just watched JoVE's introduction to the glovebox and chemical sensors. You should now understand how to operate a glovebox, how to test for water and oxygen contamination, and how to synthesize air- and moisture sensitive compounds. Thanks for watching!