Fig. 1.
Multiple-reaction monitoring LC-MS/MS chromatograms of blank (A) and fortified milk samples (B) at the level of 0.15 µg kg−1. The navy blue trace (the trace peaking directly below the given value 3.92 min) corresponds to chloramphenicol (CAP), while the pink trace (the trace extending higher) corresponds to the internal standard (CAP-D5 at the level of 1.5 µg kg−1)
Fig. 2.
Multiple-reaction monitoring LC-MS/MS chromatograms of blank (A) and fortified muscle samples (B) at the level of 0.15 µg kg−1. The navy blue trace (the trace peaking directly below the given value 3.92 min) corresponds to chloramphenicol (CAP), while the pink trace (the trace extending higher) corresponds to the internal standard (CAP-D5 at the level of 1.5 µg kg−1)
Table 1.
Validation results for chloramphenicol determination in milk and muscle matrices
| Parameter | Matrix | 0.10 µg kg−1 | 0.15 µg kg−1 | 0.30 µg kg−1 |
|---|---|---|---|---|
| RSDr (%)* | Milk | 7.5 | 6.8 | 5.6 |
| RSDwR (%)* | Milk | 8.9 | 8.1 | 6.7 |
| Apparent recovery (%) | Milk | 99.2 | 97.5 | 101.8 |
| Expanded uncertainty (µg kg−1, k = 2) | Milk | 0.021 | 0.027 | 0.035 |
| CCα (µg kg−1)* | Milk | 0.10 | - | - |
| CCβ (µg kg−1)* | Milk | 0.12 | - | - |
| RSDr (%)* | Muscle | 8.2 | 7.4 | 6.1 |
| RSDwR (%)* | Muscle | 9.8 | 8.7 | 7.2 |
| Apparent recovery (%) | Muscle | 97.8 | 98.5 | 101.2 |
| Expanded uncertainty (µg kg−1, k = 2) | Muscle | 0.028 | 0.035 | 0.042 |
| CCα (µg kg−1)* | Muscle | 0.10 | - | - |
| CCβ (µg kg−1)* | Muscle | 0.12 | - | - |
Table 2.
Effect of membrane type on micro-scale solid-phase extraction clean-up performance
| Membrane | Qualitative observations | Impact on method performance |
|---|---|---|
| PVDF | Stable flow, reproducible elution, no apparent analyte loss | Consistent analytical response and good reproducibility |
| Nylon | Inconsistent flow and partial analyte retention | Increased variability and reduced reproducibility |
| PTFE | Poor elution efficiency and unstable behaviour during centrifugation | Reduced robustness and potential analyte loss |
Table 3.
Robustness evaluation of selected micro-scale solid-phase extraction parameters
| Parameter | Tested range | Effect on analytical performance |
|---|---|---|
| Loading solvent (MeOH %) | 25–35% | No significant effect observed |
| Elution solvent (MeOH %) | 60–70% | Minor effect on analytical response |
| Elution volume | 0.18–0.22 mL | No significant effect on quantification |
| Reconstitution volume | 0.30–0.40 mL | Stable analytical response |
| Mixing time | 5–15 min | No significant differences observed |
| Membrane type | PVDF / Nylon / PTFE | PVDF provided the most stable performance |
Fig. 3.
Distribution of matrix effect values obtained for milk and muscle samples following the hybrid QuEChERS–micro-scale solid-phase extraction procedure. Dots show results for one blank sample each, boxes indicate the interquartile range, horizontal lines represent the median, and whiskers indicate the range of observed values
Table 4.
Comparison of selected LC-MS/MS methods for chloramphenicol determination in milk and muscle matrices
| Reference | Matrix | Extraction / Clean-up strategy | Analytical performance | Main characteristics |
|---|---|---|---|---|
| 14 | Bovine milk | SPE (C18) + alumina clean-up | Recovery ~90%, RSD 2.8 –4.3%, CCβ 0.03 µg/kg | SPE and alumina-based clean-up workflow |
| 10 | Multiple food and biological matrices | ACN extraction + chloroform clean-up | RSD < 25%, CCβ 0.03 –0.04 µg/kg | Multi-matrix LC-MS/MS workflow |
| 13 | Liquid milk, milk powder, bovine muscle | Ethyl acetate extraction with additional clean-up | Accuracy 89–107%, RSD 4–15% | Simultaneous CAP and florfenicol determination |
| 12 | Multiple food matrices including milk and muscle | QuEChERS | Recovery 86–111%, RSDr < 11.2% | Simplified QuEChERS workflow |
| Milk and muscle | QuEChERS + PVDF-MSPE microspin clean-up | Recovery 97.5–101.8%, RSDr < 10% | Additional microspin clean-up and matrix effect control |