Chemometric guided isolation of new triterpenoid saponins as acetylcholinesterase inhibitors from seeds of Achyranthes bidentata Blume.

Achyranthes bidentata Blume (Amaranthaceae) is an annual or perennial herb widely used as ethnomedicine in Traditional Chinese Medicine for treating fever, cold, ulcers, mensural pain, dementia, and osteoporosis. In the current study, UPLC-IM-Q-TOF-MS/MS-based chemometric approach was adopted for the tentative identification of fifty-six compounds in the extract and fractions of A.bidentata seeds. Further, the chemometric-guided isolation led to the isolation of two previously undescribed oleanane-type triterpenoid saponins, named achyranosides A-B (27 and 30), along with three known compounds (31, 44, and 23) from water fraction of A. bidentata seeds. The structures of new compounds were elucidated based on the detailed analysis of NMR, HR-ESI-MS, FT-IR spectral data, and GC-FID techniques. The isolated compounds in vitro acetylcholinesterase inhibitory activity revealed the promising activity of chikusetsusaponin IVa (23) (IC50 = 63.7 µM) with mixed type of AChE inhibition in enzyme kinetic studies. Additionally, in silico binding free energy of isolated compounds disclosed the greater stability of enzyme-ligand complex owing to underlying multiple H-bond interactions. Overall, the study demonstrates the effectiveness of a chemometric-guided approach for the phytochemical exploration and isolation of new oleanane-type triterpenoid saponins from A. bidentata seeds.

Antiplasmodial activity of the bulbs of Fritillaria cirrhosa D.Don (Syn: Fritillaria roylei Hook.): UPLC-IM-Q-TOF-MS/MS-based biochemometric approach for the identification of marker compounds.

ETHNOPHARMACOLOGICAL RELEVANCE: Fritillaria cirrhosa D.Don (Syn: Fritillaria roylei Hook.) (Hindi name: Kshirakakoli) is a critically endangered Himalayan medicinal plant, well documented in Ayurveda for its therapeutic uses against various disorders such as jvara (fever), kasa (respiratory tract disease) etc. Its bulbs are also used as Szechuan-Pei-Mu for their antipyretic properties in the traditional Chinese medicine. However, despite its ethnomedicinal usage, the therapeutic use of F. cirrhosa bulbs for jvara (fever) related conditions such as malaria has remained unexplored. Hence in the context of increasing global concerns about drug-resistant malaria, it is important to investigate the antiplasmodial activity of F. cirrhosa bulbs for novel antimalarial agents. AIM OF THE STUDY: To investigate the antiplasmodial effects of the extracts/fractions of F. cirrhosa bulbs by the biochemometric approach and to rationalize its ethnopharmacological usage for jvara (fever) related conditions such as malaria. MATERIAL AND METHODS: This study involves the UHPLC-MS-based plant material selection, preparation, quantification, and assessment of F. cirrhosa bulb extracts against CQ-sensitive Pf 3D7 & CQ-resistant Pf INDO strains. Further, UPLC-IM-Q-TOF-MS-based biochemometric approach has been applied for the identification of marker compounds responsible for the observed antiplasmodial effects. The identified marker compounds were also assessed for their in silico ADMET properties and binding efficacy with the drug transporter Pf CRT. RESULTS: Different F. cirrhosa bulb extracts/fractions showed promising antiplasmodial activity with IC50 values 2.71-19.77 µg/mL for CQ-resistant Pf INDO strain and 1.76-21.52 µg/mL for CQ-sensitive Pf 3D7 strain. UPLC-IM-Q-TOF-MS/MS-based biochemometric analysis revealed four marker compounds i.e., peimine (m/z 432.3448), peimisine (m/z 428.3504), puqiedinone (m/z 414.3379), and puqiedine (m/z 416.3509) responsible for the observed antiplasmodial activity. The identified marker compounds showed excellent binding efficacy with Pf CRT and suitable drug-like properties in silico. CONCLUSIONS: The study demonstrated promising antiplasmodial activity of the chloroform and alkaloid enriched fractions of F. cirrhosa bulbs and further identified the four marker compounds responsible for the promising antiplasmodial activity. These marker compounds i.e., peimine, peimisine, puqiedinone and puqiedine were identified by the biochemometric analysis as the putative antiplasmodial constituents of the F. cirrhosa bulbs. Further, in silico studies indicated the good binding affinity of the marker compounds with Pf CRT along with suitable ADMET properties. Overall, the study elucidates the antiplasmodial activity of F. cirrhosa bulbs from the western Himalayan region and provides nascent scientific evidence for their ethnopharmacological usage in jvara (fever) related conditions such as malaria.

A novel extraction method enhanced the osteogenic and anti-osteoporosis effect of tea extract without any hepatotoxicity in ovariectomized rats.

Tea (Camellia sinensis) has several reported health benefits, including that on bone health attributed to catechins of which the most abundant is epigallocatechin-3-gallate (EGCG). However, several preclinical and clinical studies raise safety concerns about EGCG in tea extract causing acute liver failure. Tea also contains kaempferol, albeit scanty, and it has hepatoprotective and osteogenic effects. Here, we utilized a novel extraction procedure of acid hydrolysis to enhance the osteogenic effect of tea extract while reducing its hepatotoxicity. The resultant extract (USKECSE) has a ~40-fold increase in kaempferol and a 2.5-fold reduction in EGCG content compared with the hydroethanolic extract (USCSE). In a female Sprague Dawley (SD) rat femur osteotomy model, USKECSE (100 mg/kg) but not USCSE promoted bone regeneration. In a rat postmenopausal osteoporosis model induced by bilateral ovariectomy (OVX), USKECSE through an osteogenic mechanism maintained bone mass, strength, and microarchitecture to the levels of ovary-intact rats with no hepatotoxic effect. After a single oral dose (100 mg/kg) of USKECSE to adult rats, kaempferol was detectable for 48 hours, suggesting its significant absorption and distribution in plasma. Peak kaempferol concentration in plasma (Cmax) was 483 ng/ml (2 µM), and at this concentration, kaempferol induces osteoblast differentiation. USKECSE had no genotoxicity, and its safety index assessed by preclinical toxicity studies, including safety pharmacology, was >20-fold. Taken together, we report a novel extraction process that enhanced the osteogenicity and concomitantly reduced hepatotoxicity of tea extract with significant kaempferol bioavailability and a favorable systemic safety profile. Based on these data, we propose assessing the USKECSE effect for postmenopausal osteoporosis treatment.

Govanoside B, a new steroidal saponin from rhizomes of Trillium govanianum.

Trillium govanianum, commonly known as Nag Chhatri and Teen Patra, is a popular herbal supplement traditionally used for curing different inflammatory and sexual disorders, infection and wound healing. Steroidal saponins are considered as active components of this species. The present study demonstrated the isolation of nine steroidal saponins, including one new compound named as govanoside B (9) and eight known, pregna-chacotrioside (1), pennogenin-triglycoside (2), borassoside E (3), pennogenin-tetraglycoside (4), protodioscin (5), clintonioside B (6), pennogenin-diglycoside (7) and borassoside D (8). This is the first report on the isolation of 1, 2, 4, 5, 6, 7 and 8 from rhizomes of T. govanianum. The extract, fractions and isolated compounds were further evaluated for their DPPH and ABTS radical scavenging activity.

Qualitative and quantitative determination of steroidal saponins in Trillium govanianum by UHPLC-QTOF-MS/MS and UHPLC-ELSD.

INTRODUCTION: Trillium govanianum (Nag Chhatri and Teen Patra) is traditionally used for curing joint pains, wounds, and sexual disorders. Steroidal saponins are the main active components of this species. However, only a small amount of information is available about steroidal saponins of this plant. OBJECTIVE: To develop an ultra-high-performance liquid chromatography-quadrupole time of flight tandem mass spectrometry (UHPLC-QTOF-MS/MS) and ultra-high-performance liquid chromatography-evaporative light scattering detector (UHPLC-ELSD) methods for the qualitative and quantitative determination of steroidal saponins in T. govanianum. METHOD: The dried rhizomes of T. govanianum (100 mg) were extracted with ethanol-water (80:20, 10 mL) by ultrasonic treatment for 30 min at 40°C. The prepared sample was analysed by UHPLC-QTOF-MS/MS and UHPLC-ELSD for the qualitative and quantitative determination of steroidal saponins. RESULT: A total of 24 saponins were identified using UHPLC-QTOF-MS/MS; seven of them were characterised by comparing with standards. Furthermore, five saponins [govanoside B (2), protodioscin (6), pennogenin tetraglycosides (11), borassoside E (21) and borassoside D (24)] were quantified using UHPLC-ELSD method in different extracts and fractions of T. govanianum. The method showed good linearity (R2 ≥ 0.993), limit of detection (0.92-4.09 µg/mL), limit of quantification (3.1-13.5 µg/mL), precision [intra-day relative standard deviations (RSDs) < 4.3% and inter-day RSDs < 5.5%], and accuracy (84.0-110.3%). This is the first report on the quantification of 2, 6, 11, 21 and 24 in T. govanianum. CONCLUSION: The present study provides an efficient analytical method for the identification and quantification of steroidal saponins and will be helpful for the quality evaluation of T. govanianum.