In vivo metabolic tracking of ¹⁴C-radiolabelled isoflavones in kudzu (Pueraria lobata) and red clover (Trifolium pratense) extracts

Abstract

Absorption, distribution and elimination of 14C-labelled isoflavone-containing extracts from kudzu (Pueraria lobata) root culture and red clover (Trifolium pratense) cell culture were investigated in an in vivo rat model. The predominant isoflavones in the kudzu extract were the glycosides puerarin, daidzin and malonyl daidzin, while in the red clover extract, the major isoflavones were formononetin and its derivatives, genistein and biochanin A, with radioactivities of 3.770 and 7.256 MBq/g, respectively. Male Sprague-Dawley rats, implanted with a jugular catheter and a subcutaneous ultrafiltrate probe, were orally administered with 14C-labelled isoflavone extracts from either kudzu or clover cell cultures. Serum, interstitial fluid (ISF), urine and faeces were collected using a Culex Automated Blood Collection System for 24 h. Analysis of bone tissues revealed that radiolabel accumulated in the femur, tibia and vertebrae at 0.04, 0.03 and 0.01 % of the administered dose, respectively, in both kudzu and red clover treatments. The liver accumulated the greatest concentration of radiolabel among the tissues tested, at 1.99 and 1.54 % of the administered kudzu and red clover extracts, respectively. Serum and ISF analysis showed that both extracts were rapidly absorbed, distributed in various tissues, and largely eliminated in the urine and faeces. Urine and faeces contained 8.53 and 9.06 % of the kudzu dose, respectively, and 3.60 and 5.64 % of the red clover dose, respectively. Serum pharmacokinetics suggest that extracts from kudzu may undergo enterohepatic circulation.

Tracking deposition of a 14C-radiolabeled kudzu hairy root-derived isoflavone-rich fraction into bone

“…Isoflavone aglycones, for instance, are absorbed faster and in higher amounts than their glucosides. 40 A plateau in serum concentration at four hours after gavage, which followed the trend seen in the metabolic tracking of 14 C-kudzu crude extract, 27 is consistent with the enterohepatic circulation of kudzu isoflavones. 41 Plasma collected 24 h after gavage showed no difference in 14 C activity compared with serum collected 24 h after gavage.…”

“…27 This may be attributed to the removal of non-target phytochemicals from the crude extract, including sugars, which would also have acquired radiolabel, and would have been readily absorbed and distributed throughout all animal tissues. Rats gavaged with the isoflavone-rich kudzu fraction accumulated 0.011%, 0.09% and 0.003% of the administered dose in each femur, tibia and vertebrae, respectively (Table 4).…”

“…Previously, we showed that 14 C-labeled phytochemicals can be isolated from kudzu (P. lobata) roots as a crude extract, identified, quantified 26 and tracked in vivo to evaluate absorption, distribution into organs and clearance from the body. 27 In this study, we have significantly optimized the radiolabeling system by: (1) screening kudzu germplasm to select an elite, high-puerarin-yielding genotype; (2) inducing hairy roots by inoculation with Agrobacterium rhizogenes to further magnify isoflavone accumulation; 28 and (3) purifying an isoflavone-enriched radiolabeled subfraction from the crude extract, which was then tracked in vivo to determine bone accumulation. 30 Gamborg B5 vitamins, 31 0.1 g/L myoinositol (Phytotechnology Laboratories), 30 g/L sucrose, 0.1% FeEDTA and 2 g/L gellan gum (PhytoTechnology Laboratories).…”

Neutral Loss Ion Mapping Experiment Combined with Precursor Mass List and Dynamic Exclusion for Screening Unstable Malonyl Glucoside Conjugates

“…Some minor malonyl glycosides, such as malonyl daidzin and malonyl genistin, were also reported from Gegen in the past decades [19][20][21]. To our knowledge, there were no C-glycoside malonyl conjugates reported till now.…”

Puerarin stimulates osteoblasts differentiation and bone formation through estrogen receptor, p38 MAPK, and Wnt/β-catenin pathways

“…Puerarin was able to protect neurons against the toxicity of 1-methyl-4-phenylpyridinium (MPP þ )-induced apoptosis and protect myocardium against ischemia and reperfusion injury [9,10]. By using 14 C radiolabeled analysis, puerarin had been shown to accumulate in bone tissues of rats, the femur, tibia, and vertebrae at 0.04%, 0.03%, and 0.01% of the oral administered dose, respectively [11]. A previous study also reported that puerarin promoted bone formation through activation of phosphatidylinositol-3-kinase/AKT pathway [3].…”