Honey, as a commercial product, is a target of adulteration through inappropriate production practices and deliberate mislabelling of botanical origin. Floral nectar protein could be a good marker for determining the source flowers of honey, especially monofloral honeys. Here, nectar and monofloral honey from Eriobotrya japonica Lindl. (loquat) were systematically compared, especially regarding proteomic and enzymatic activity. Using two-dimensional electrophoresis and mass spectrometry, only bee-originated proteins were detected in loquat honey. Xylosidase, thaumatin, and two kinds of chitinases were detected in loquat floral nectar, and their activity in loquat nectar and honey were quantified. Following gel electrophoresis, loquat honey had similar chitinase activity profiles to loquat nectar, but both were clearly distinguishable from Camellia sinensis nectar and Brassica napus honey. To our knowledge, this is the first examination of nectar-origin enzyme activity in honey. Zymography of chitinases is a potential marker for determining or authenticating the botanical origin of honeys.

Chitin is a linear homo-polymer of N-acetyl-D-glucosamine (GlcNAc) and the second most abundant biopolymer after cellulose. Several industries rely on the bioprocesses for waste chitin recycle and hydrolysis by chitinase (EC 3.2.1.14) for potential healthcare applications through the production of its monomeric subunit, GlcNAc. In the present study, a chitinase-producing fungus (named as MFSRK-S42) was isolated from the marine water sample of North Bay of the Andaman and Nicobar Islands. It was identified as Aspergillus terreus by morphological and molecular characterization methods leveraging the internal transcribed spacer between 18S rRNA and 5.8S rRNA. Chitinase that was isolated from the fermentation broth of marine Aspergillus terreus was used to carry out biotransformation of chitineaceous wastes. Prior to the enzymatic hydrolysis step, chitins from different sources were characterized for the presence of characteristic functional groups, grain size distribution, and surface morphology. Enzymatic hydrolysis of 50 mg/ml substrate with six units of enzyme incubated for 5 days revealed 15, 36.5, 40, and 46 mg/ml GlcNAc production from ground prawn shell, chitin flakes, colloidal prawn shell, and swollen chitin respectively under standardized conditions, as determined by HPLC. In this study, 30, 73, 80, and 92% GlcNAc yields were observed from ground prawn shell, chitin flakes, colloidal prawn shell, and swollen chitin conversion respectively. The HPLC-eluted product was confirmed as GlcNAc by the presence of characteristic functional groups in FTIR and 244 Da molecular weight peak in HRMS analyses.

Kinetin, a cytokinin which promotes seed germination by inhibiting the action of abscisic acid, is an important molecule known to trigger various molecular mechanisms by interacting with an array of proteins shown from experimental observations in various model organisms. We report here the prediction of most probable protein targets of kinetin from spinach proteome using in silico approaches. Inverse docking and ligand-based similarity search was performed using kinetin as molecule. The former method prioritized six spinach proteins, whereas the latter method provided a list of protein targets retrieved from several model organisms. The most probable protein targets were selected by comparing the rank list of docking and ligand similarity methods. Both of these methods prioritized chitinase as the most probable protein target (ΔG pred = 5.064 kcal/mol) supported by the experimental structure of yeast chitinase 1 complex with kinetin (PDB: 2UY5) and Gliocladium roseum chitinase complex with 3,7-dihydro-1,3,7-trimethyl-1H-purine-2,6-dione (caffeine; 3G6M) which bears a 3D similarity of 0.43 with kinetin. An in vitro study to evaluate the effect of kinetin on spinach seed germination indicated that a very low concentration of kinetin (0.5 mg/l) did not show a significant effect compared to control in inducing seed germination process. Further, higher levels of kinetin (>0.5 mg/l) constituted an antagonist effect on spinach seed germination. It is anticipated that kinetin may have a molecular interaction with prioritized protein targets synthesized during the seed germination process and reduces growth. Thus, it appears that kinetin may not be a suitable hormone for enhancing spinach seed germination in vitro.