Effects of Royal Jelly Freshness and Concentration on Lifespan, Growth, Motility and Reproduction in Caenorhabditis elegans

Although aging is an irreversible process, the rate of aging can be delayed by a reasonable diet. As a nutrient-dense natural product, royal jelly (RJ) has an enormous potential for applications in medicine and health promotion. However, the exact physiological activity of RJ with varying freshness and concentration has not been fully clarified, and more investigation is needed to determine their precise contributions. Here, fresh RJ (just produced recently) and RJ stored for 2 weeks at -20 °C, 4 °C or 25 °C were tested at concentrations of 100, 50, 25 and 12.5 μg/mL on <i>Caenorhabditis elegans</i>. Fresh RJ, with concentrations of 100 μg/mL, 50 μg/mL and 25 μg/mL, could extend the lifespan of <i>C. elegans</i> by 16.37%, 9.53% and 4.32%, while RJs stored at 4 °C and 25 °C were ineffective. In terms of body length, treatment with fresh RJ significantly enlarged the body size by around 48%. Although RJ stored at 4 °C and 25 °C could also promote nematode growth, its activity diminishes as storage temperature increases. RJs stored at -20 °C and 4 °C with concentrations of 100 μg/mL significantly increased the pumping rate of nematodes by 58% and 50%. But non-fresh RJ or low-concentration RJ (≤25 μg/mL) had no effects on the motility of <i>C. elegans</i>. In addition, fresh RJ could improve the reproductive capacity of <i>C. elegans</i>, with the highest increase reaching approximately 25%. Even when stored at 25 °C, RJ also significantly enhanced the reproductive capacity of <i>C. elegans</i>, increasing it by approximately 14.8%. Moreover, qPCR showed that RJ could significantly affect the expression of multiple genes associated with aging and vitality. Fresh RJ significantly up-regulated <i>bec1</i> and <i>hsp16.2</i> 3.19- and 2.80-fold, while RJ stored at 25 °C significantly up-regulated <i>sod3</i> and <i>gpd1</i> 3.80- and 3.40-fold. Our results suggested that the activity of RJ on <i>C. elegans</i> is related to its freshness and concentration, while RJ also contains active components that are independent of freshness. Therefore, it is necessary to explore effective methods for accurately assessing the freshness of RJ.