Local Climate and Cultivation Practice Shape Total Protein and Phenolic Content of Mulberry (Morus sp.) Leaves in Sub-Mediterranean and Sub-Pannonian Regions of Slovenia

Mulberry (Morus sp.) trees, traditionally cultivated for their leaves used in sericulture, have recently gained recognition for their adaptability and valuable ecosystem services. The biochemical composition of mulberry leaves varies both qualitatively and quantitatively, depending on genotype, environmental conditions, and cultivation practices. This study aimed to (1) identify differences in old local white (M. alba L.) and black mulberry (M. nigra L.) leaves, (2) perform a chemotype analysis of monitored local varieties, and (3) evaluate the influence of selected bioclimatic factors and pruning practices on the biochemical composition of leaves of white mulberry trees across Slovenian mesoregions. Black mulberry exhibited a higher phenolic content, particularly caffeoylquinic acid derivatives (16.05 mg/g dry weight (DW)), while white mulberry contained more quercetin glycosides (6.04 mg/g DW). Ward’s clustering identified three chemotypes, two of which had elevated protein and hydroxycinnamic acid levels, making them particularly suitable for silkworm feeding. Considering pruning practices of white mulberries, we determined significantly increased protein contents in yearly pruned trees (187.24 mg/g DW). Principal component analysis revealed interactions between bioclimatic, morphological, and biochemical factors, distinctly separating mulberries from the Sub-Mediterranean and Sub-Pannonian macroregions. White mulberries from Sub-Pannonian regions accumulated more caffeoylquinic acids in leaves under lower precipitation and total insolation, while those from Sub-Mediterranean regions exhibited higher kaempferol derivatives due to photo-thermal stress. These findings highlight the influence of climate and pruning on mulberry biochemical diversity and adaptation.