2.2.4. Recovery

The tests were performed by using three different concentrations to test the recovery values in the linearity range of the method.

Quantities of CBD (4, 8, and 24 μg/mL) were added, thus assessing concentrations similar to, higher, and lower than those found in samples.

Recovery was determined according to this modality for CBD and was 84.92%.

An evaluation of recovery on all the compounds present in the sample was carried out by proceeding with a further extraction with 10 mL of methanol-chloroform on the sample residue after the usual extraction; in this extract, some cannabinoids were present, and indirectly the percentage of recovery was determined.


= (10 × σ)/m). 3 LOD determined by the signal-to-noise ratio (Instrumental LOD: S/N = 3). 4 LOQ determined by the signal-to-noise ratio (Instrumental LOQ: S/N = 10). \* Not detectable.

**Table 1.** Validation parameters of RP-HPLC-UV method.

5

The percentage of recovery values, as shown in Table 1, were higher than 84.92% and can be considered very satisfactory. In fact, considering CBD, the percentages are higher than those previously reported in the literature [5].

#### 2.2.5. Detection Limit, LOD

The instrumental limit of detection was determined by the calibration curve, according to the formulas expressed in Section 3.6. The instrumental limit of detection (LOD) values obtained for CBDA and CBGA (Table 1) were lower, while those of CBG and CBD were comparable with similar methods described in literature [5,27]. Low LOD values were found also for the other cannabinoids (THCV, CBN, Δ-9 THC, Δ-8 THC, CBC, THCA), indicating that the method is sensitive.

#### 2.2.6. Quantification Limit, LOQ

The instrumental limit of quantification was determined by a calibration curve, according to the formulas expressed in Section 3.6, considering that the signal-to-noise method is particularly useful to quantify the cannabinoids present at lower concentrations, such as THC. As reported for the LODs, the instrumental limit of quantification (LOQ) values obtained for CBDA and CBGA (Table 1) were also lower than those reported in the literature, while those for CBG and CBD were comparable with those of other methods described for similar procedures [5,27]. In addition, the other cannabinoids (THCV, CBN, Δ-9 THC, Δ-8 THC, CBC, THCA) showed low LOQs. The instrumental noise was registered in μV, by performing 3 blank injections with the ASTM method [28] given by the instrument, and a maximum CV% of 3.49% was calculated for all individual compounds to determine the single LOD and LOQ, which was considered acceptable.
