Vehicle-Controlled In Vitro Screening of a Pilot-Scale Mandarin (Citrus reticulata L.) Peel Extract Against Monilinia sp. and Rhizopus stolonifer

Mandarin (Citrus reticulata L.) peel is generated in appreciable quantities as a processing residue in the northern Ecuadorian highlands, and its pilot-scale ethanolic extract has already been developed and compositionally documented by our group as a botanical insecticide. The present study extends that valorisation cascade to postharvest crop protection, and in doing so addresses a methodological problem that pervades the by-product bioactivity literature: the contribution of a crude extract is seldom separated from that of the surface-active adjuvant used to disperse it. An extract produced by the established protocol from peel of documented provenance (d-limonene confirmed in the previous experiments as the principal detected constituent by APGC-MS; extraction yield 9.97% w/w) was assayed at 2.5% (v/v) against Monilinia sp. and Rhizopus stolonifer by the poisoned-medium technique. Colonised area was measured every 24 h from standardised photographs using ImageJ within a completely randomised, vehicle-controlled design (n = 3 dishes per treatment), and analysed by a linear mixed-effects model on log-transformed area with dish as the repeated experimental unit. Relative to untreated medium, the complete formulation inhibited growth by 35.2% (Monilinia sp.) and 37.3% (R. stolonifer); the organosilicone adjuvant alone, however, reproduced 22.7% and 29.1% of that effect respectively, and no consistent incremental contribution of the extract over the vehicle could be demonstrated for either pathogen. The study therefore yields a transferable result for by-product valorisation research: inhibition percentages reported for crude extracts in the absence of a vehicle control are not attributable to the extract, the attribution error here amounting to between roughly two-thirds and four-fifths of the recorded effect.