## 5 Safety and Bioavailability of Nicotinamide Riboside
### Nicotinamide Riboside Bioavailability  
In mice and humans, NR is the most orally bioavailable of NAD\({}^{+}\) precursors. Amounts of NAD\({}^{+}\) in mouse liver were elevated by oral NRS with better pharmacokinetics (PK) than seen with NA and NAM. In one healthy volunteer, a pilot study showed that a single dose of oral NRS elevated serum NAD\({}^{+}\) levels as much as 2.7-fold. The first clinical trial of NR PK in healthy humans highlighted that 100, 300, and 1000 mg of NR single dose increased the NAD\({}^{+}\) metabolome in a dose-dependent manner [231].  
An open-label, non-randomized study of the PK of the NR nutritional supplement and its effects on blood NAD\({}^{+}\) levels was performed in eight healthy volunteers. Oral NR (250 mg) was administered daily for the first two days. Then, the maximum dose was increased to 1000 mg twice daily on days 7 and 8. NRS did not induce adverse effects and was well tolerated. In NR and NAD\({}^{+}\) levels, starting from baseline to the ninth day, the absolute changes were highly correlated [232; 233].  
BM stromal cell antigen 1 (BST1) regulates NR metabolism via its dual roles in glyco-hydrolase and base-exchange enzyme activities. Two different pathways contribute to the increase in NAD\({}^{+}\) level after oral NR treatment: (i) NR salvage pathway in the early phase of NR direct absorption, and (ii) Preiss-Handler pathway in the late phase of NR hydrolysis to NAM by BST1 followed by transformation to NA by the distal gut microbiome [234; 235].  
Pre-steady-state and steady-state kinetic analyses were made for the \(N\)-ribosyl hydrolase activity of hCD157. Cell-surface hCD157 binds and catalyzes the slow hydrolysis of NR and NA riboside (NAR) after the formation of an initial complex with NAD\({}^{+}\) derivatives [236].  
The import of NR and NAR into human cells is mediated by the equilibrative nucleoside transporter (ENT) family of proteins, ENT1, ENT2, and ENT4. After entering the HEK93 cells, NR is activated by NRK phosphorylation in the cytosol and then transformed to yield NAM [237]. NMN and NR metabolism in mammalian cells is predominantly controlled by NRK1 and purine nucleoside phosphorylase (PNP). Simultaneous PNP downregulation could improve the health advantages of NRS [14; 238].  
[PERSON] et al. (2015) studied the generation, release, and uptake of the NAD\({}^{+}\) precursor NAR by human cells. Under normal culture conditions, untransfected HeLa cells generated and released a sufficient level of NAR and NR as whole compounds derived from NAD\({}^{+}\) metabolism [239].  
A pilot study investigated changes in the human plasma and urine NAD\({}^{+}\) metabolome during a 6 h i.v. infusion of 3 uM/min NAD\({}^{+}\). No changes were seen after two hours in the levels of plasma NAD\({}^{+}\) or its metabolites, such as NAM, MeNAM, adenosine diphosphate ribose (ADPR), and NMN. After six hours, increased urinary excretion of NAD\({}^{+}\) and MeNAM was seen; at the same time, NAM urinary level was not increased [240].  
In a simulated intestinal fluid using porcine pancreatic, NR trioleate chloride, a newly synthesized hydrophobic NR derivative, was partly digestible and easily released NR [241].  
NRH is a potent enhancer of NAD\({}^{+}\) levels in vitro in mammalian cells, significantly increasing NAD\({}^{+}\)/NADH ratio, as well as in vivo after i.p. injection to C57 BL/6] mice [242].  
Although there are minor structural dissimilarities, NRH defines a new pathway for NAD\({}^{+}\) biosynthesis, independent of NRK1, and acts as an orally bioavailable NAD\({}^{+}\) precursor also in mammalian cells and mouse tissues [243]. In a recent study, it was found that NR and dihydronicitmic acid riboside synergistically act to increase NAD\({}^{+}\) intracellular levels by generating NRH [244].