**4. Conclusions**

This work proposes the sampling of gunshot residues on shooters' hands using dry cotton swabs followed by vortex-assisted extraction with water over a short time (20 s). Aqueous samples were directly processed in the miniaturized IT-SPME-CapLC-DAD system for on-line clean-up and preconcentration of the sample and for quantization of the amount of diphenylamine as targeted organic residue. It is worth mentioning that non-toxic solvents and low-cost materials were employed. The efficiencies of the IT-SPME were tested for several compositions and lengths of the extractive phase, as well as sample volume processed in order to improve the sensitivity. The highest analytical responses were obtained for the longest TRB-35 capillaries (90 cm) were more likely due to π–π interactions and 1.8 mL of processed volume. The proposed approach is a rapid, green, and cost-effective option for detecting DPA on the hands of shooters. The sustainability of an analytical method is governed by minimization of toxic solvents, reduction of wastes, and employment of energy-efficient and cost-effective methodologies, but also on maintaining the reliability of the performance parameters, such as sensitivity, precision, and accuracy [25,26]. In two previous papers [27,28] our group demonstrated that IT-SPME-CapLC-DAD achieves the minimization of the sample pre-treatment step, analysis time, and wastes, the reduction of the analysis costs, and thus, improvement of the analytical and environmental performance. Satisfactory LOD (0.3 ng) and precision (RSD intra-day = 9%, RSD inter-day = 14%) were achieved.

In order to test the utility of the method for real cases, several shooters' hands were sampled by dry cotton swabs and processed by IT-SPME-CapLC-DAD. The results showed that DPA was found and quantified in 81% of samples. Additionally, IGSRs inspection of swab samples was carried out by optical microscopy in order to confirm the presence of gunshot residues on shooters' hands, which were analyzed by SEM-EDS after DPA extraction. Furthermore, some shooters' hands were sampled by a tape lift kit, which is the typical police sampler, but DPA extraction was fourteen times lesser than that achieved by the dry cotton swab sampler. Morphology, elemental composition, and distribution of the IGSRs particles were also studied. Then, improved results were obtained by the proposed sampling method as indicated above. If organic compounds are detected in combination with inorganic compounds, higher probative value can be achieved, and false positives/negatives can also be reduced for discriminating shooters' hands. In this work, a sensitive chromatographic method to detect the organic compound DPA can be combined with IGSR analysis by SEM-EDS in order to obtain valuable evidence of GSRs deposited on hands of a suspected shooter. Therefore, the proposed method is helpful to determine whether a person has fired a gun in a forensic investigation.

**Author Contributions:** All authors designed and performed part of the experiments, analyzed the data, and wrote the paper.

**Acknowledgments:** The authors are grateful to the EU-FEDER/MINCIU-AEI of Spain (project CTQ2017-90082-P) and Generalitat Valenciana (PROMETEO 2016/109) for the financial support received and to Police Headquarters of Valencian Community (Valencia, Spain) for the sampling.

**Conflicts of Interest:** The authors declare no conflict of interest. The founding sponsors had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, and in the decision to publish the results.
