Method validation is fundamental for ensuring the accuracy, reliability, and regulatory compliance of analytical data in pharmaceuticals, #QualityControl, R&D, and natural product analysis.

1. #Specificity
How to Perform: Inject blank, placebo, standard, and sample solutions.
What to Evaluate: Confirm there's no interference at the analyte’s retention time.
Applicable Instruments: HPLC, LC-MS, GC, etc.

2. #Linearity
How to Perform: Prepare at least five concentrations (e.g., 80%, 90%, 100%, 110%, 120%).
What to Evaluate: Plot concentration vs. peak area, and calculate the correlation coefficient (R² ≥ 0.995).
Applicable Instruments: HPLC, UV, etc.

3. #Accuracy / #Recovery
How to Perform: Spike known amounts of standard into the sample at three levels (e.g., 80%, 100%, 120%).
What to Evaluate: %Recovery = (Detected / Added) × 100
Acceptance Range: 98–102% for APIs
Replicates: Three per level

4. #Precision
Repeatability (Intra-day): Analyze six replicates of the same concentration on the same day.
Intermediate Precision (Inter-day): Repeat on different days, using different analysts or instruments.
What to Evaluate: %RSD should be ≤ 2%

5. Limit of Detection (LOD)
How to Perform: Use the formula LOD = 3.3 × (σ/S), where σ = standard deviation of the response, and S = slope.
Alternative Approach: Use a signal-to-noise ratio of 3:1.

✔️ 6. #LimitOfQuantitation / #LOQ
How to Perform: LOQ = 10 × (σ/S)
Alternative Approach: Signal-to-noise ratio of 10:1
More stringent than LOD

7. #Range
How to Determine: Based on results from linearity, accuracy, and precision studies.
What to Evaluate: Must cover 80–120% of the target analyte concentration.

⚙️ 8. #Robustness
How to Perform: Slightly vary method parameters:
• Mobile phase pH ± 0.2
• Flow rate ± 0.1 mL/min
• Column temperature ± 2°C
What to Evaluate: Ensure %RSD and retention time remain within acceptable limits.

9. #Ruggedness
How to Perform: Conduct analysis under varying conditions:
• Different analysts
• Different instruments
• Different laboratories
What to Evaluate: Results should remain consistent (%RSD ≤ 2%)

✅ 10. System Suitability
How to Perform: Inject system suitability solution before and during the analysis.
Key Parameters:
• %RSD (area or RT) ≤ 2%
• Theoretical plates (N) ≥ 2000
• Tailing factor ≤ 2
• Resolution ≥ 2 between critical peaks

These validation parameters are the foundation of reliable results in #HPLC, LC-MS, and other analytical techniques.

Are all these checkpoints part of your method validation workflow?

#QualityControl #MethodValidation #AnalyticalScience #ICHQ2 #PharmaTesting #MethodVerification #RegulatoryCompliance