“The assay value of the drug and total responses of all degradants should equal the initial value i.e. it’s 100% is mass balance”.
Mass balance in pharmaceutical analysis is very important and is used all over the pharmaceutical industry. In the synthetic development, the mass balance calculation is used to assess efficiency and yields. It represents the degradative losses of the parent drugs compared well with the measured increase in the degradation products. Equally, if one observes, for example, a 10% loss of parent drug but only measures a 2 % increase in degradation products, it is expected that additional degradation products formed are not accurately determined by the present method(s).
Why Mass balance?
Mass balance is an important consideration in assessing the degradation pathways of pharmaceutical products. The related substances or organic impurities method is a stability-indicating method for degradants. it is important to have methods that detect all major degradation products. Thus, safety is the major reason for the study of mass balance.
Mass balance is also useful in method validation. In order to demonstrate that analytical methods are stability-indicating, unstressed, and stressed results were compared and evaluated. Any increase in degradation of a product that correlates well with the loss of parent drug aids in demonstrating that the methods can accurately assess degradation.
How much Degradation?
Usually, 5 % to 10 % of degradation is looked for as a forced degradation study during the different forced degradative conditions. Not all compounds/formulations will degrade by 10% under even extreme conditions, however, a sufficient attempt must be taken to promote degradation. Any stability observed from the studies must be scientifically explained and justified in case of low degradation levels (i.e. < 10%). These types of criteria can be used within internal guidance documents where extreme conditions are defined.
Why Mass Balance Not Achieved?
During Stress studies it is “good to show the mass balance” but in reality, it is normally not achieved due to the following reasons,
- Forced degradation is to prove the selectivity of the method in terms of interference to the main peak and major known impurities.
- Since the conditions used are very harsh and not usually encountered in real stability studies. Such stress conditions generate secondary and tertiary degradation.
- It will never know whether the resultant products are UV active or not, in certain cases, gases liberated from the sample, and the end product might be acid with no UV chromophore or just precipitation.
- The degradant formed may not be eluted in the developed method (extend runtime or modify method conditions).
- The Relative Response Factor may be zero at the method wavelength (select the appropriate wavelength or develop a separate method).
Figure: Fishbone diagram of the major causes of mass imbalance.
How Do We Achieve Mass Balance?
The primary aim is to separate related substance degradants from unrelated, ones to identify key degradation pathways by studying chemistry to get MS and UV spectra on degradants. Develop a simple method to resolve the degradants from the drug and from each other. Calculating the mass balance from this initial study tells us how much we know.
We need to explore different detectors like GC/MS, LC/MS, LC-MS-NMR, RI Detector, and ELSD Detector for possible degradants. Later steps might include the isolation of specific degradants to determine structure and response factors, giving us a better understanding of chemistry and a quantitative correction to the degradant’s response.
How is Mass Balance Calculated?
There are no such guidelines on acceptance criteria for Mass Balance; however, the industry practice is to prove the Mass Balance results in the range of 95%-105% during Related Substance Analytical Method Validation. It is the basic prerequisite to state your method is Stability Indicating in addition to the rest validation parameters. The above statement on Forced Degradation studies is acceptable provided the Mass Balance is achieved in the above prescribed range.
The mass balance equation:
Total mass = (Amount of mass remained + Amount of known degradants + Amount of unknown degradants)
So, here will convert the equation for better understanding. After each degradation study,
Total % of drug = % drug remained + % known degradants + % unknown degradants
- Relative response factors play an important role in calculating the exact mass balance.
- The molecular weight of Drugs and the molecular weight of the degradant formed play an important role in calculating the exact mass balance.
- International Conference on Harmonization (ICH) and US Food and Drug Administration, Q1B: Photostability Testing of New Drug Substances and Products.
- ICH and FDA, Q1A(R2): Stability Testing of New Drug Substances and Products.
- ICH and FDA, Q2A: Validation of Analytical Procedures, Text for Validation of Analytical Procedure.
- Impurities in new drug substances, Q3A, Q3B, and Q3C, ICH of Technical Requirements For Registration of Pharmaceuticals.
- ICH and FDA, Q2B: Validation of Analytical Procedures, Methodology.
- M. Nussbaum et al., “Determination of Relative UV Response Factors for HPLC by use of a Chemiluminescent Nitrogen-Specific Detector,” J. Pharm. Biomed.
- Baertschi, S.W., Alsante, K.M., Reed, R.A. (Eds.). (2011). Pharmaceutical Stress Testing: Predicting Drug Degradation (2nd ed.). London, UK: Informa Healthcare.