Glycerol Units Research Articles

Biocatalytic Ether Lipid Synthesis by an Archaeal Glycerolprenylase

AbstractAlthough ethers are common in secondary natural products, they are an underrepresented functional group in primary metabolism. As such, there are comparably few enzymes capable of constructing ether bonds in a general fashion. However, such enzymes are highly sought after for synthetic applications as they typically operate with higher regioselectivity and under milder conditions than traditional organochemical approaches. To expand the repertoire of well characterized ether synthases, we herein report on a promiscuous archaeal prenyltransferase from the scarcely researched family of geranylgeranylglyceryl phosphate synthases (GGGPSs or G3PSs). We show that the ultrastable Archaeoglobus fulgidus G3PS makes various (E)‐ and (Z)‐configured prenyl glycerol ethers from the corresponding pyrophosphates while exerting perfect control over the configuration at the glycerol unit. Based on experimental and computational data, we propose a mechanism for this enzyme which involves an intermediary prenyl carbocation equivalent. As such, this study provides the fundamental understanding and methods to introduce G3PSs into the biocatalytic alkylation toolbox.

Biocatalytic Ether Lipid Synthesis by an Archaeal Glycerolprenylase.

Although ethers are common in secondary natural products, they are an underrepresented functional group in primary metabolism. As such, there are comparably few enzymes capable of constructing ether bonds in a general fashion. However, such enzymes are highly sought after for synthetic applications as they typically operate with higher regioselectivity and under milder conditions than traditional organochemical approaches. To expand the repertoire of well characterized ether synthases, we herein report on a promiscuous archaeal prenyltransferase from the scarcely researched family of geranylgeranylglyceryl phosphate synthases (GGGPSs or G3PSs). We show that the ultrastable Archaeoglobus fulgidus G3PS makes various (E)- and (Z)-configured prenyl glycerol ethers from the corresponding pyrophosphates while exerting perfect control over the configuration at the glycerol unit. Based on experimental and computational data, we propose a mechanism for this enzyme which involves an intermediary prenyl carbocation equivalent. As such, this study provides the fundamental understanding and methods to introduce G3PSs into the biocatalytic alkylation toolbox.

Phosphate-Containing Glycolipids: A Review on Synthesis and Bioactivity.

Phosphate-containing glycolipids (PcGL) are scarcer than the better understood glycolipids. They are composed of arrangements of phosphate, carbohydrates and glycerol units and are always found associated with lipids. PcGL are often found associated with cell membranes, suggesting they play roles in cell membrane structure and intercellular interactions. This article aims to provide an up-to-date overview of the existing knowledge and research on PcGL, emphasizing their synthesis and wide range of biological activities. When it comes to the synthesis of PcGL compounds, the strategies for glycosylation mainly rely on the thioglycoside donor, the trichloroacetamidate donor and halide donor strategies, while phosphorylation is stapled and falls on either phosphite chemistry or phosphoryl chloride chemistry. Certain bacteria utilize PcGLs in their pathogenicity, triggering an inflammatory response within the host's defense mechanisms. The best-known examples of these structures are teichoic acids, lipopolysaccharide and the capsular polysaccharide found in bacteria, all of which are frequently implicated in bacterial infections. Given the degree of variability within PcGL structures, they were found to display a wide range of bioactivities. PcGL compounds were found to: (1) have anti-metastatic properties, (2) behave as agonists or antagonists of platelet aggregation, (3) be mostly pro-inflammatory, (4) display antifungal and antibiotic activity and (5) have neurogenic activity.

Synthesis of Prepolymers of Poly(glycerol-co-diacids) Based on Sebacic and Succinic Acid Mixtures.

In this study, poly(glycerol-co-diacids) prepolymers were produced using different ratios of glycerol (G), sebacic acid (S), and succinic acid (Su) (molar ratios: GS 1:1, GSSu 1:0.9:0.1, GSSu 1:0.8:0.2, GSSu 1:0.5:0.5, GSSu 1:0.2:0.8, GSSu 1:0.1:0.9, GSu 1:1). All polycondensation reactions were performed at 150 °C until reaching a degree of polymerization of ≈55%, inferred by the water volume collected from a reactor. We concluded that the reaction time is correlated with the ratio of diacids used, that is, the increase in succinic acid is proportional to a decrease in the duration of the reaction. In fact, the reaction of poly(glycerol sebacate) (PGS 1:1) is twice as slow as the reaction of poly(glycerol succinate) (PGSu 1:1). The obtained prepolymers were analyzed by electrospray ionization mass spectrometry (ESI-MS) and 1H and 13C nuclear magnetic resonance (NMR). Besides its catalytic influence in poly(glycerol)/ether bond formation, the presence of succinic acid also contributes to a mass growth of ester oligomers, the formation of cyclic structures, a greater number of oligomers detected, and a difference in mass distribution. When compared with PGS (1:1), and even at lower ratios, the prepolymers produced with succinic acid presented mass peak characteristics of oligomer species with a glycerol unit as its end group in higher abundance. Generally, the most abundant oligomers have molecular weights between 400 and 800 g/mol.

Thermal decomposition mechanism investigation of hyperbranched polyglycerols by TGA-FTIR-GC/MS techniques and ReaxFF reactive molecular dynamics simulations

Molecular structure of hyperbranched polyglycerols (hbPGs) strongly affected the thermal stability as the critical heat transfer medium to maintain thermal homeostasis during working of heat engine. Here, thermogravimetric analysis (TGA)-fourier-transform infrared spectroscopy (FTIR)-gas chromatography (GC)-mass spectroscopy (MS) and reactive force-field (ReaxFF) techniques were utilized to identify major thermal decomposition products and explore the molecular mechanism. TGA-FTIR-GC/MS results showed that the major products were CO, CO2, water molecules and C3 organic moieties. To perform ReaxFF molecular dynamic simulation, timestep-dependent images revealed that C3 molecules were mainly originated from glycerol units and hydroxyl groups played an essential role in catalyzing thermal decomposition. To examine our hypothesis, different methoxylation levels of hbPGs were prepared, and TGA analysis of these methoxylated hbPGs polymers clearly corroborated that lower percentage of hydroxyl groups in the hbPGs can significantly increase maximum peak temperature from ∼400 °C to ∼460 °C. Our investigation clearly provided the molecular mechanism of hbPGs thermal decomposition and identified a potential way to improve thermal stability of hbPGs.

Thiamyxins: Structure and Biosynthesis of Myxobacterial RNA-Virus Inhibitors.

During our search for novel myxobacterial natural products, we discovered the thiamyxins: thiazole- and thiazoline-rich non-ribosomal peptide-polyketide hybrids with potent antiviral activity. We isolated four congeners of this unprecedented natural product family with the non-cyclized thiamyxin D fused to a glycerol unit at the C-terminus. Alongside their structure elucidation, we present a concise biosynthesis model based on biosynthetic gene cluster analysis and isotopically labelled precursor feeding. We report incorporation of a 2-(hydroxymethyl)-4-methylpent-3-enoic acid moiety by a GCN5-related N-acetyltransferase-like decarboxylase domain featuring polyketide synthase. The thiamyxins show potent inhibition of RNA viruses in cell culture models of corona, zika and dengue virus infection. Their potency up to a half maximal inhibitory concentration of 560 nM combined with milder cytotoxic effects on human cell lines indicate the potential for further development of the thiamyxins.

Lipase-Catalyzed Poly(glycerol-1,8-octanediol-sebacate): Biomaterial Engineering by Combining Compositional and Crosslinking Variables.

This study examined poly(glycerol-1,8-octanediol-sebacate) (PGOS) copolymers with low-level substitution of O (1,8-octanediol) for G (glycerol) units (G/O ratios 0.5:0.5, 0.66:0.33, 0.75:0.25, 0.8:0.2, and 0.91:0.09) prepared in bulk by immobilized Candida antarctica Lipase B (N435) catalysis. The central question explored was the extent that exchanging less than half of poly(glycerol sebacate) (PGS) glycerol units with 1,8-octanediol can be used as a strategy to fine-tune biomaterial properties. Synthesized copolymers having G/O ratios of 0.66:0.33, 0.75:0.25, 0.8:0.2, and 0.91:0.09 have similar molecular weights, where Mw varied from 52,800 to 63,800 g/mol, Mn varied from 5100 to 6450 g/mol, and ĐM (molecular mass dispersity, Mw/Mn) values were also similar (8.4-11.4). All of the copolymers were branched, and dendritic glycerol units reached 11% for PGOS-0.91:0.09:1.0. Analysis of DSC second heating scans revealed that copolymers with higher 1,8-octanediol contents have relatively higher Tm and ΔHf values. Over the copolymer compositional range studied herein, Tm and ΔHf values varied from 5.3 to 21.1 °C and 8.0 to 23.1 J/g, respectively. Stress-strain curves of PGOS copolymers cured at 140 °C for 48 h exhibited either a unimodal, bimodal, or trimodal response to tensile loading. Varying G/O from 10:1 to 2:1 resulted in significant increases in the peak stress (0.26-4.01 MPa), preyield modulus (0.65-62.59 MPa), failure to strain (64-110%), and failure toughness (0.1-0.56 MPa). This demonstrates that altering the G/O ratio over a narrow compositional range provides biomaterials with widely different yet tunable mechanical properties. Further investigation of PGOS-0.75:0.25:1.0 films revealed that varying the cure conditions from 120 to 160 °C for periods of 24-72 h provides access to biomaterials with a failure strain range from 15 to 224% and Young's modulus from 1.17 to 10.85 MPa. Hence, using the dual variables of compositional variation and changes in cure conditions provides an exciting platform for PGS analogues to optimize material-tissue interactions. Increased contents of 1,8-octanediol slowed in vitro degradation. Slowed degradation of PGOS relative to PGS will be valuable for use in slower healing wounds.

Eco-conception of Highly Salt-Tolerant Alkyl Ether Carboxylate Hydrotropes with a Glyceryl Spacer.

New alkyl ether carboxylates with a glyceryl spacer instead of ethylene glycol units have been synthesised using environmentally friendly methodology. A cascade synthesis of acetalisation and hydrogenolysis was developed to obtain products containing an alkyl chain linked to a glycerol unit bearing a polar carboxylate head. These products were methylated by using trimethyl phosphate to observe the influence of a free or methoxylated alcohol on the physicochemical properties. Finally, saponification gave the carboxylate anionic group of the new hydrotropes. Studying the amphiphilicity, the tolerance to sodium and calcium ions, and the solubilising power of these bio-based ionic/nonionic hydrotropes has shown that they exhibit significantly improved application properties compared to similar petro-based hydrotropes.

Lipase TL®-mediated kinetic resolution of glycerol analogues: Efficient convergent route to both enantiomeric glycerol units

In the presence of pyridine, at −5 °C, lipase TL®-mediated kinetic resolution of (±)-1-((tert-butyldimethylsilyl)oxy)-3-((4-methoxybenzyl)oxy)-propan-2-ol was found to proceed efficiently to give the corresponding (S)-alcohol and (R)-acetate, each with high enantiomeric excess and in quantitative yield. The alcohol (S)-1 and the acetate (R)-2-derived alcohol, (R)-1, could be converted to the 1-protected glycerols with either of the stereochemistry by the choice of the deprotection of the protective groups.

Highly Sensitive Detection of CB [7] Based on Fluorescence Resonance Energy Transfer between RhB and Gold Nanoparticles

Background: Cucurbit[n]uril (CB[n], n=5, 6, 7, 8 and 10）is a type of macrocyclic compounds formed by n glycerol units and 2n methylene. The different sizes of the cavity lead to the different chemical characteristics of CB[n]. Therefore, it is very important to distinguish and detect CB[n]. Methods: At room temperature, Rhodamine B was added to a gold nanoparticles solution. The above mixture was respectively mixed with different concentrations of Cucurbituril[n] or β-cyclodextrin solutions. The fluorescence spectra and ultraviolet-visible spectra of samples were determined. Results: It was found that 1.2nM gold nanoparticles could quench the fluorescence of 1μM Rhodamine B completely. After adding a certain amount of Cucurbituril[7], the fluorescence intensity of Rhodamine B was restored. Compared with other macrocyclic compounds, such as Cucurbituril[5], Cucurbituril[6] and β-cyclodextrin, we found that this method had unique selectivity for Cucurbituril[ 7]. Under the optimal conditions, the fluorescence recovery efficiency was linearly proportional to the concentration of Cucurbituril[7] in the range of 0.8-8 μg·mL-1. The detection limit was 0.21 μg·mL-1. Conclusion: The research established an effective and practical FRET-based detection method for CB[7] with RhB as a donor and the gold nanoparticles as the acceptor. The system had unique and extensive selectivity for CB[7].

Synthesis of Nucleoside Derivatives of N-Acetyl-7-nitroindoline, Their Incorporation into the DNA Oligomer, and Evaluation of Their Photoreactivity in the DNA/RNA Duplex.

N-Acetyl-7-nitroindoline has a characteristic reaction in that its acetyl group is photo-activated to acetylate amines to form amides. In this study, the N-acetyl-7-nitroindoline part was connected to the 2'-deoxyribose part at the 3- or 5-position or to a glycerol unit at the 3-position through an ethylene linker (1, 2, and 3, respectively). They were incorporated into the oligodeoxynucleotides, and their photo-reactivities toward the complementary RNA were evaluated. The acetyl group of 1 was photo-activated to form the deacelylated nitroso derivative without affecting the RNA strand. The photoreaction with 2 suggested acetylation of the RNA strand. In contrast, compound 3 formed the photo-cross-linked adduct with the RNA. These results have shown the potential application of N-acetyl-7-nitroindoline unit in aqueous solutions.

Enzymatic Polymerization of Poly(glycerol-1,8-octanediol-sebacate): Versatile Poly(glycerol sebacate) Analogues that Form Monocomponent Biodegradable Fiber Scaffolds.

A family of poly(glycerol sebacate) (PGS) analogues were synthesized by Candida antarctica lipase B (CALB) catalysis to tailor biomaterial properties. Different fractions of glycerol (G) units in PGS were replaced by 1,8-octanediol (O) units. Poly(glycerol-1,8-octanediol-sebacate), PGOS, synthesized by CALB catalysis with a 1:3 molar ratio of G to O units has Mn and Mw values of 9500 and 92,000, respectively. PGS undergoes fiber fusion during electrospinning, and cross-linked PGS rapidly resorbs when implanted. By decreasing the molar ratio of glycerol-to-octanediol from 1:1 to 1:4, the peak melting temperature (Tm) increased from 27 to 47 °C. PGOS with 1:3 G to O units was electrospun into nanofibers without the need for a second component. The copolymer is semicrystalline and, when cross-linked, undergoes slow in vitro mass loss (3.5 ± 1.0% in 31 days) at pH 7.4 and 37 °C. Furthermore, PGOS cross-linked films have an elastic modulus of 106.1 ± 18.6 MPa, which is more than 100 times that of cross-linked PGS. New PGOS polymers showed tunable molecular weights, better thermal properties, and excellent electrospinnability. This work expanded PGS analogues' function, making these suitable biodegradable polymers for various biomedical applications.

Evaluation of nanomechanical properties of hyperbranched polyglycerols as prospective cell membrane engineering block

Owing to the excellent biocompatibility, hyperbranched polyglycerols (hbPGs) are one of the most promising polymers and widely employed in drug delivery. Presented as an excellent bioinert coating material, hbPGs can significantly improve the biosafety of biomedical nanomaterials. However, it is still unclear what specific properties of hbPGs are the key effectors to bioinertness. Here, atomic force microscopy was employed to test the Young’s modulus and adhesion of hbPGs, spin-coated onto mica substrate. High Young’s modulus indicated that the hbPGs cannot be further compressed and low adhesion implied that it is not easy to form hbPGs aggregators. This could owe to the intramolecular hydrogen bond. Morphology characterization of hbPGs self-assembled monolayer onto Si(100) substrate, confirmed the lower adhesion among different hbPGs and indicated their biofouling properties. Further confocal laser microscopy of cell membrane modified with alkyl chain (C18)-modified hbPGs and hbPGs-NH2, confirmed that the antifouling properties of hbPGs are determined by terminal glycerol units. Our findings demonstrated that only hbPGs with entire terminal surface can be used as perspective cell membrane modification skeleton.

Chemical characterisation of polymerised extractives in bleached birch kraft pulp

Abstract The main aim of this study was to chemically characterise the polymeric fraction of birch pulp extractives. We showed that 70–96% of the material in extracts of fully bleached birch kraft elemental chlorine-free (ECF) pulp from three Finnish mills consists of compounds that are undetectable by conventional gas chromatography (GC) analysis, i.e. high-molar mass (HM) material. There were small variations in the extractive content and composition between the three mills, but the overall trend was the same. The HM material was present mainly in the acetone extracts, and the molar mass was shown to range between ca. 1000 and 10 000 Da. Alkaline hydrolysis of the hexane and acetone extracts released fatty acids (FAs) especially, but also sterols and triterpenoids. The dominating FAs were palmitic and stearic acids. Pyrolysis and thermally assisted hydrolysis and methylation (THM)-GC-mass spectrometry (MS) analyses confirmed that the HM material consists mainly of FA units, and additionally of sterol, triterpenoid and aromatic units. Nuclear magnetic resonance (NMR) analysis further confirmed the presence of FAs containing aromatic groups, but also glycerol units. The FA chains seemed to be linked to each other by ester linkages mainly, although ether linkages were also present. The FA oligomers and polymers may have a suberin-like structure.

Design and synthesis of ethoxylated esters derived from waste frying oil as anti-ultraviolet and efficient primary plasticizers for poly(vinyl chloride)

In this work, a series of novel bio-based plasticizers, including ethoxylated esters, methylated esters, and glycidyl esters, originated from waste frying oil (WFO), were prepared, characterized, and used as primary plasticizers for Poly (vinyl chloride) (PVC). The glass transition temperature, mechanical properties, transparency, UV resistance, thermal stability and migration resistance of plasticized PVC were investigated and compared with that of the PVC sample plasticized by the commercial dioctyl terephthalate (DOTP). It is found that all of those properties are significantly depended on the structures of ester groups. In particular, the esters bearing ethyoxyl units show better plasticization than that having methyl or glycidyl groups. In addition, the plasticizing efficiencies of WFO-derived ethoxylated esters are proportional to the number of ethyoxyl units in their structures. Among them, the anti-ultraviolet PVC sample plasticized by WFO-derived polyacid (triethylene glycol monomethyl ether) esters (WFOPA-3, “3” refers to the number of ethylene glycerol units in the feedstock of triethylene glycol monomethyl ether) with maximum ethyoxyl units exhibited the best plasticization, thermal stability, and transparency than the properties of other samples plasticized by WFO-based plasticizers, as well as petro-based DOTP. The results suggest WFOPA-3 is an attractive potential as alternative to DOTP.

A method for the quantitative determination of glycerophospholipid regioisomers by UPLC-ESI-MS/MS

Diacyl glycerophospholipids (GPs) belong to the most abundant lipid species in living organisms and consist of a glycerol backbone with fatty acyl groups in sn-1 and sn-2 and a polar head group in the sn-3 position. Regioisomeric mixed diacyl GPs have the same fatty acyl composition but differ in their allocation to sn-1 or sn-2 of the glycerol unit. In-depth analysis of regioisomeric mixed diacyl GP species composed of fatty acyl moieties that are similar in length and degree of saturation typically requires either chemical derivatization or sophisticated analytical instrumentation, since these types of regioisomers are not well resolved under standard ultra-performance liquid chromatography (UPLC) conditions. Here, we introduce a simple and fast method for diacyl GP regioisomer analysis employing UPLC tandem mass spectrometry (MS/MS). This GP regioisomer analysis is based both on minor chromatographic retention time shifts and on major differences in relative abundances of the two fatty acyl anion fragments observed in MS/MS. To monitor these differences with optimal precision, MS/MS spectra are recorded continuously over the UPLC elution profile of the lipid species of interest. Quantification of relative abundances of the regioisomers was performed by algorithms that we have developed for this purpose. The method was applied to commercially available mixed diacyl GP standards and to total lipid extracts of Escherichia coli (E. coli) and bovine liver. To validate our results, we determined regioisomeric ratios of phosphatidylcholine (PC) standards using phospholipase A2-specific release of fatty acids from the sn-2 position of the glycerol backbone. Our results show that most analyzed mixed diacyl GPs of biological origin exhibit significantly higher regioisomeric purity than synthetic lipid standards. In summary, this method can be implemented in routine LC-MS/MS-based lipidomics workflows without the necessity for additional chemical additives, derivatizations, or instrumentation.

Glycerol-modified poly(ε-caprolactone): an biocatalytic approach to improve the hydrophilicity of poly(ε-caprolactone)

Novel poly(e-caprolactone) (PCL)–glycerol polyester was synthesized in one step in the presence of Novozym 435 under mild reaction conditions. The objective of the study was to investigate the feasibility of enhancing the hydrophilicity of PCL by condensing it with glycerol through biocatalytic approach. Effect of reaction time, glycerol-to-CL ratio and CL-to-toluene ratio was studied. The 1H NMR and GPC results established that self-condensation of CL units and condensation of CL with glycerol units take place simultaneously. When the glycerol content was 10 and 20 mol% with regard to CL, condensation continued for 6 h; further extension of reaction time resulted in the cleavage of ester linkages. When the glycerol content was increased to 40 mol%, the condensation of CL–glycerol units was accomplished in 4 h; however, the reaction had to get extended beyond 6 h for complete condensation of CL–CL units. The study revealed that the crystallinity and the melting temperature (Tm) of the products altered with an increase in the glycerol content. Furthermore, TGA and contact angle values of water on PCL and its glycerol derivative films revealed that the water uptake capacity of PCL too increased significantly with the increase in glycerol content, rendering PCL more hydrophilic.

Synthesis, Characterization and 3D Micro-Structuring via 2-Photon Polymerization of Poly(glycerol sebacate)-Methacrylate–An Elastomeric Degradable Polymer

Poly(glycerol sebacate) (PGS) has been utilised in numerous biomaterial applications over recent years. This elastomeric and rapidly degradable polymer is cytocompatible and suited to various applications in soft tissue engineering and drug delivery. Although PGS is simple to synthesise as an insoluble prepolymer, it requires the application of high temperatures for extended periods of time to produce an insoluble matrix. This places limitations on the processing capabilities of PGS and its possible applications. Here, we present a photocurable form of PGS with improved processing capabilities: PGS-methacrylate (PGS-M). By methacrylating the secondary hydroxyl groups of the glycerol units in the PGS prepolymer chains, the material was rendered photocurable and, in combination with a photoinitiator, crosslinked rapidly on exposure to UV light at ambient temperatures. The polymer’s molecular weight and the degree of methacrylation could be controlled independently and the mechanical properties of the crosslinked material tailored. The polymer also displayed rapid degradation under physiological conditions and cytocompatibility with various primary cell types. As a demonstration of the processing capabilities of PGS-M, µm scale 3D scaffold structures were fabricated using 2-photon polymerisation and used for 3D cell culture. The tunable properties of PGS-M coupled with its enhanced processing capabilities make the polymer an attractive potential biomaterial for various future applications.

Synthesis of Alkyl-Glycerolipids Standards for Gas Chromatography Analysis: Application for Chimera and Shark Liver Oils.

Natural O-alkyl-glycerolipids, also known as alkyl-ether-lipids (AEL), feature a long fatty alkyl chain linked to the glycerol unit by an ether bond. AEL are ubiquitously found in different tissues but, are abundant in shark liver oil, breast milk, red blood cells, blood plasma, and bone marrow. Only a few AEL are commercially available, while many others with saturated or mono-unsaturated alkyl chains of variable length are not available. These compounds are, however, necessary as standards for analytical methods. Here, we investigated different reported procedures and we adapted some of them to prepare a series of 1-O-alkyl-glycerols featuring mainly saturated alkyl chains of various lengths (14:0, 16:0, 17:0, 19:0, 20:0, 22:0) and two monounsaturated chains (16:1, 18:1). All of these standards were fully characterized by NMR and GC-MS. Finally, we used these standards to identify the AEL subtypes in shark and chimera liver oils. The distribution of the identified AEL were: 14:0 (20–24%), 16:0 (42–54%) and 18:1 (6–16%) and, to a lesser extent, (0.2–2%) for each of the following: 16:1, 17:0, 18:0, and 20:0. These standards open the possibilities to identify AEL subtypes in tumours and compare their composition to those of non-tumour tissues.

8,4′-Oxyneolignane glucosides from an aqueous extract of “ban lan gen” (Isatis indigotica root) and their absolute configurations

Three pairs of glycosidic 8,4′-oxyneolignane diastereoisomers, named isatioxyneolignosides A−F (1–6), were isolated from an aqueous extract of Isatis indigotica roots. Their structures and absolute configurations were elucidated by comprehensive spectroscopic data analysis and enzyme hydrolysis. The validity of ΔδC8-C7 values to distinguish threo and erythro aryl glycerol units and Cotton effects at 235±5nm to determine absolute configurations at C-8 in 1–6 and their aglycones (1a–6a) are discussed.