Stage Three: Designing the Sample Preparation
The third stage in constructing E/L studies is designing the sample preparation to achieve the AET, and the possible conversion
of extracts to appropriate solvents for instrumental analysis. Liquid to liquid extraction is often used to make solvents
more appropriate for GC analysis (volatiles and semivolatiles). Extracts may also be concentrated by evaporation for improved
sensitivity, or, for solvent exchange, dried and reconstituted in an appropriate solvent for the analysis.
Stage Four: Analyzing Extracts, and Evaluating and Compiling Data
The next stage involves analyzing material solvent extracts, and evaluating and compiling data to identify extractable compounds.
Multiple analytical techniques are typically used to fully profile the material extract solutions. Headspace GC/MS is used
to analyze the extracts for volatile compounds, direct liquid injection GC/MS for semivolatiles, and LC/MS for nonvolatiles
(typically by both atmospheric pressure chemical ionization [APCI] positive and negative modes of detection). If applicable,
metals screening is performed by ICP/MS. Compounds of special concern to the FDA because of their carcinogenicity, such as
formaldehydes, polyaromatic hydrocarbons, mercaptobenzothiozole, and nitrosamines, are analyzed using specialized instrumentation
and detection, sample derivatization, or other techniques. The headspace and direct injection GC/MS extractables data are
evaluated by a library compound software search, and these data, as well as the LC/MS spectral pattern data, are reviewed
and evaluated for tentative compound identification. Tentative peak confirmations and semi-quantitative amounts typically
are confirmed by obtaining commercial standards of the known extractable compounds (if available), and by injecting these
compounds to confirm retention times and spectral patterns.
A toxicological evaluation of extractables data is critical in determining which compounds need to be tracked as potential
leachables in the drug product. Typically, TDI values calculated for the confirmed, known extractable compounds are provided
by the analytical laboratory to the board-certified reviewing toxicologist. The toxicologist evaluates the compounds and amounts
observed for toxicological concern or structure alert. The resulting information, along with knowledge of the intended use
of the product, is applied when deciding which of the identified extractable compounds should be tracked as potential leachables
in the drug product, and what extractable compounds should be tracked in routine methods for raw material quality control
screening (quality control release methods for vendor-supplied materials).
Methods are developed next for monitoring leachables. The methods are selective for identified leachables and for expected,
unidentified leachables. The methods separate these peaks from drug product or process impurity chromatographic peaks. The
leachable methods are developed to provide sufficient sensitivity, accuracy, precision, and linearity across a suitable range,
to quantitatively determine the amount of leachable components present. Because these procedures are for routine, quality
control (QC) use, they typically use high-performance liquid chromatography/ultraviolet visible (HPLC/UV) and gas chromatography/flame
ionization detector (GC/FID) techniques, as opposed to mass spectrometric detection.
At this stage, it is also advisable to develop methods for monitoring extractables in incoming vendor raw material batches
before these are used in the build process for a final product. To develop the most efficient process, the results of the
controlled extraction studies should be used to optimize the routine extraction procedures applied to the incoming materials
CASE STUDY 1: EXTRACTABLES AND LEACHABLES STUDY DESIGN FOR DISPOSABLE BAGS
Disposal bags intended for bioprocessing must be qualified for extractables and leachables before use to support manufacturing
process qualification and validation. Data collected for the respective materials also can be used to compile a knowledge
base and enhance scientific understanding which, in preparation for a future paradigm shift into quality by design per ICH
Q8 (pharmaceutical development), will support ICH Q9 (quality risk management) and ICH Q10 (pharmaceutical quality systems).
The entire extractables and leachables qualification for a disposable bag is described here for a one-liter solution disposable
bag with a two-doses-per-day regimen.
Step 1: Information Gathering (Component selection and final formulation determination). An appropriate bag should be chosen for the intended storage and
use of the disposable drug product. The properties of the material the bag is made of (such as pH tolerance, thermal resistance,
tensile strength, and other related chemical and physical properties), and the processes and additives used in the bag-making
process, including how the plastics polymer is made, are crucial information to gather.
The final formulation and the bioprocess conditions (holding time, temperature, pH, etc.) are gathered when they are determined.
For example: the drug placebo is a saline solution; the bags hold 1 L of the drug solution for at most 3 months; the bags
are processed at 35 °C; and the dosage for the final drug product is 1 mL/dose and 2 doses/day.