Native stingless bees (NSBs) are widely distributed in Brazilian territory. They are responsible for pollinating plants, maintaining biodiversity, and producing honey, a natural and therapeutic sweetener, used by different populations. This study aimed to identify physical, chemical, and microbiological characteristics of honey samples collected during the first harvesting of Tetragonisca angustula, Melipona mondury, and Scaptotrigona postica bees kept in the first public meliponary situated in Marília, São Paulo state, Brazil. Samples were collected directly from NSB’s hives in the meliponary and sequentially matured in plastic bottles with airlock valves, duringn 90 days, in the Physicochemical Analyzes Laboratory at Fatec Marília. Subsequently, analyzes were carried out to determine pathogenic and spoilage microorganisms. In addition, analyzes to determine free acidity, reducing sugars, total soluble solids, apparent sucrose, humidity, insoluble solids, pH, water activity, and hydroxymethylfurfural were performed. Results showed the presence of molds and yeasts. Nevertheless, because the values agreed with the current legislation and the pathogenic and deteriorating contaminants were absent in the honey samples, one may suggest that they were brought to the hive by bees, since floral nectaries are their normal habitats. The physicochemical parameters, with the exception of reducing sugars, agreed with other scientific studies and reinforced the proposal for technical regulation, considering the absence of specific national legislation for NSB’s honey. Therefore, it is considered relevant to establish regulations for NSB’s honey to define identity and standard requirements to guarantee safe consumption and direction for scientific studies in the area.