Neither scheme of Method A extracted as much water from the samples as Method C. The difference is more pronounced for high-moisture
samples (see the circles in Figure 1), where Method C also yielded less scatter in the data. However, Method C may be influenced
by the level of relative humidity in the room because the crushed lyophilized cake is briefly exposed to ambient conditions.
Hence, the true moisture content of a sample may lie between the values given by Methods A and C.
The authors attempted to estimate the tendency for moisture uptake during sample preparation for Method C by leaving uncapped
vials on the bench for 2–3 min after the contents were crushed. The difference in moisture content between these dwelled samples
and ones that were analyzed immediately after pulverization was typically less than 0.1% (w/w). It is not possible to know
the absolute amount of environmental moisture introduced during the entire sequence of operation, but Method C tends to err
on the safer side because it may overestimate samples' water content.
Although Method A offers the advantage of protecting the sample from direct handling and exposure to ambient conditions, the
additional steps required after extraction are undesirable. Furthermore, products contained in large containers such as 50–100-cm3 vials will require longer extraction time because the water has a longer diffusion path. Method A also requires large volumes
of solvent for the extraction, thus compounding waste-disposal problems. For these reasons, the authors decided to implement
Method C in the automated system.
The authors also considered fully automating sample preparation, including uncapping, crushing, and dispensing of the lyophilized
cake. Such a system, however, would need to handle vials sizes from 5 to 100 cm3 and to consistently pulverize lyophilized product cake of various consistencies and various heights. These capabilities
would entail a complex and cost-prohibitive setup for sample preparation alone. Hence, the authors simply elected to prepare
samples manually, as is current practice. A trained analyst can perform these operations in less than 2 min per sample.
Automated-system design and construction
Sequence of operation.
Given the requirements discussed above, the automated system should operate according to the following sequence:
- The analyst prepares the sample according to Method C. An amount (e.g., 50–150 mg) of crushed lyophilizate is dispensed into
a standard centrifuge tube, which is then tightly capped. The tube is placed into a sample rack. Several samples can be prepared
and loaded into the system.
- The automated system retrieves a sample tube from the rack and transfers it to a balance that records the weight.
- The sample tube is placed in a decapping station, and the tube cap is removed.
- The sample is dispensed into the titration cell. Titration is initiated.
- The sample tube is recapped and reweighed. The actual sample mass delivered to the titration cell is determined, and the sample
tube is discarded.
- The titration result is returned, and the titration cell is cleaned if necessary after a prespecified number of titrations.
- The cycle repeats from step 2.
This sequence is basically the same as that of the manual technique. A robotic system is needed to shuttle the sample between
the rack, balance, and titrator. The titration of the sample remains the rate-limiting step, so the total analysis time is
comparable to that of a fully manual operation. Titration times typically range from a few minutes to more than 10 min, depending
on the composition and moisture level of the sample. The benefit of an automated setup, however, is the ability to perform
unattended analysis once the samples have been loaded into the system.
To minimize costs, the authors incorporated as many off-the-shelf components as possible. A Mettler-Toledo XP 204 balance
was selected because this model offers RS-232 serial connectivity, motorized draft doors, and a removable front control panel.
The latter feature is important because removing the front panel prevents the accidental activation of a balance operation
during a run.