# Influence of Ingestion of Lactulose on \(\gamma\)-Lactones Enanating from Human Skin Surface
## 3 Results  
Figure 3 shows a comparison between the 16S rRNA gene copy numbers (GCNs) of bifidobacteria per gram of faeces before and after 2-week ingestion of lactose powder at a dose of 4 g d\({}^{-1}\). The arithmetic mean and standard deviations are shown for six healthy participants recruited first. As can be predicted based on previous works, the common logarithm of GCNs g\({}^{-1}\) faeces significantly increased from 8.91 \(\pm\) 0.20 to 9.27 \(\pm\) 0.37 after the 2-week ingestion (Wilcoxon signed-rank test, \(p\) = 0.046). Figure 4 shows the variations in the dermal emission fluxes of C6-C11 lactones and SCFAs measured for six healthy participants from their forearms. The average emission fluxes in the morning and at night on each sampling day were used as daily values, and the arithmetic mean and standard deviation of the daily values of the six participants are shown in this figure. The dermal emission fluxes of \(\gamma\)-lactones before lactulose intake (Day 1 and 2) were in the order of C11 = C9 \(>\) C8 \(>\) C10 \(>\) C7 = C6. However, the order changed after the ingestion of lactulose powder (Day 6, 10, and 17). A remarkable increase in the cutaneous emission was observed for the C7, C10, and C11 lactones, resulting in the following order: C11 \(>\) C9 = C7 \(>\) C10 \(>\) C8 \(>\) C6. For example, the dermal emission fluxes of C11 lactone  
Figure 3: Comparison of the 16S rRNA gene copy numbers of bifidobacteria in faeces of 6 healthy participants before (Day 2) and after (Day 17) daily ingestion of lactulose for 2 weeks (\(n=6\)). at Day 1 was 1.6 \(\pm\) 0.77 ng cm\({}^{-2}\) h\({}^{-1}\), and equivalent to 1.6 \(\pm\)0.89 ng cm\({}^{-2}\) h\({}^{-1}\) at Day 2. After the ingestion of the disaccharide powder, the flux increased to 3.0 \(\pm\) 1.3 ng cm\({}^{-2}\) h\({}^{-1}\) at Day 6 (four days after the ingestion), 3.2 \(\pm\) 0.98 ng cm\({}^{-2}\) h\({}^{-1}\) at Day 10 (eight days after the ingestion), and 3.2 \(\pm\) 0.91 ng cm\({}^{-2}\) h\({}^{-1}\) at Day 17 (one day after the end of ingestion). A Jonckheere trend test was applied to the variations from Day 2 to Day 17. The results indicated significant increasing trends in the dermal emission fluxes of C7 lactone (\(p\) = 0.022) with approx. 2.8 times increase from Day 2 to Day 17, C10 lactone (\(p\) = 0.025) with approx. 1.8 times increase, and C11 lactone (\(p\) = 0.002) with approx. 2.0 times increase after oral intake of lactose powder. No apparent trend was found in the cutaneous emissions of C6, C8, and C9 lactones. The reason is not clear at present for the differences in the trends of \(\gamma\)-lactones. Figure 4: Variation in the fluxes of dermal emission of \(\gamma\)-lactones and SCFAs collected from the forearm of 6 healthy participants before (Day 1 and 2) and after (Day 6, 10, and 17) daily ingestion of lactulose. Jonckheere trend test was conducted for the values from Day 2 to Day 17. On the other hand, the variation of dermal emission was different for SCFAs. The emission flux of acetic acid gradually increased from 2.8 \(\times\) 10\({}^{2}\)\(\pm\) 52 ng cm\({}^{-2}\) h\({}^{-1}\) at Day 2 to 3.5 \(\times\) 10\({}^{2}\)\(\pm\) 1.8 \(\times\) 10\({}^{2}\) ng cm\({}^{-2}\) h\({}^{-1}\) at Day 17 (Jonckheere trend test, \(p\) = 0.09). No apparent change was observed for propionic acid, whereas a significant decrease was found in the dermal emission flux of butyric acid from Day 2 to Day 17 (\(p\) = 0.029). However, the simple sum of cutaneous emissions of these three SCFAs showed an increasing tendency after ingestion of lactulose because the acetic acid emission was much greater than other two acids. Figure 5 presents scatter diagrams of the 16S rRNA gene copy numbers of bifidobacteria in the faeces and the dermal emission fluxes of \(\gamma\)-lactones in all 10 healthy volunteers. The X-axis shows the changes in log GCNs g\({}^{-1}\) from Day 2 to Day 17, _d_log GCNs g\({}^{-1}\); the Y-axis represents the changes in dermal emissions from Day 2 to Day 17, _dE_ (ng cm\({}^{-2}\) h\({}^{-1}\)).