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CAS No. : | 6556-12-3 | MDL No. : | MFCD00166981 |
Formula : | C6H10O7 | Boiling Point : | - |
Linear Structure Formula : | - | InChI Key : | IAJILQKETJEXLJ-QTBDOELSSA-N |
M.W : | 194.14 | Pubchem ID : | 65041 |
Synonyms : |
D-(+)-Glucuronic Acid
|
Num. heavy atoms : | 13 |
Num. arom. heavy atoms : | 0 |
Fraction Csp3 : | 0.67 |
Num. rotatable bonds : | 5 |
Num. H-bond acceptors : | 7.0 |
Num. H-bond donors : | 5.0 |
Molar Refractivity : | 37.57 |
TPSA : | 135.29 Ų |
GI absorption : | Low |
BBB permeant : | No |
P-gp substrate : | No |
CYP1A2 inhibitor : | No |
CYP2C19 inhibitor : | No |
CYP2C9 inhibitor : | No |
CYP2D6 inhibitor : | No |
CYP3A4 inhibitor : | No |
Log Kp (skin permeation) : | -9.31 cm/s |
Log Po/w (iLOGP) : | -0.83 |
Log Po/w (XLOGP3) : | -2.57 |
Log Po/w (WLOGP) : | -3.29 |
Log Po/w (MLOGP) : | -3.01 |
Log Po/w (SILICOS-IT) : | -1.95 |
Consensus Log Po/w : | -2.33 |
Lipinski : | 0.0 |
Ghose : | None |
Veber : | 0.0 |
Egan : | 1.0 |
Muegge : | 2.0 |
Bioavailability Score : | 0.56 |
Log S (ESOL) : | 0.91 |
Solubility : | 1560.0 mg/ml ; 8.04 mol/l |
Class : | Highly soluble |
Log S (Ali) : | 0.27 |
Solubility : | 365.0 mg/ml ; 1.88 mol/l |
Class : | Highly soluble |
Log S (SILICOS-IT) : | 2.9 |
Solubility : | 154000.0 mg/ml ; 793.0 mol/l |
Class : | Soluble |
PAINS : | 0.0 alert |
Brenk : | 1.0 alert |
Leadlikeness : | 1.0 |
Synthetic accessibility : | 3.26 |
Signal Word: | Warning | Class: | N/A |
Precautionary Statements: | P261-P305+P351+P338 | UN#: | N/A |
Hazard Statements: | H315-H319-H335 | Packing Group: | N/A |
GHS Pictogram: |
* All experimental methods are cited from the reference, please refer to the original source for details. We do not guarantee the accuracy of the content in the reference.
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With sulfuric acid; water; sodium sulfate Electrolysis.anschl. Erwaermen mit wss. H2SO4; | ||
With platinum on activated charcoal; water; sodium hydrogencarbonate unter Einleiten von Luft und anschl. Behandeln mit wss. H2SO4; | ||
With water; nitric acid; sodium nitrite anschl. Erwaermen mir wss. H2SO4; |
With chloroform; dinitrogen tetraoxide anschl. mit wss. H2SO4; | ||
With tetrachloromethane; dinitrogen tetraoxide anschl. mit wss. Methanol und dann mit wss. H2SO4; |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
Stage #1: chitosan With hydrogenchloride In water Stage #2: D-Glucuronic acid With sodium hydroxide; 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride In water for 3h; | 1.1.7 1. 1.0 g chitosan was added to 100 mL of deionized H2O acidified with 2 mL of ION HCl.The mixture was stirred overnight to allow chitosan to go into solution.2. 1.5 g of glucuronic acid dissolved in 25 mL of deionized H20 added and the solution pH was adjusted to 4.7 with 2N NaOH3. 2.0 g of EDC was added and pH maintained at 4.7 for 3 hours.4. The solution Glu-chitosan conjugate was dialyzed extensively against deionized H20 and maintained in solution. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride; sodium hydroxide; 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride In water for 3h; | 10 Example 10 Preparation of glucopyranosyl- polvethyleneimine (Glu-PEI)1.100 mg of PEI (0.35 ml of 30% water solution) was added to 2.7 ml of IN HCl. The pH was adjusted to 4.7.2.200 mg of glucuronic acid dissolved in 1.2 ml of of deionized H20and the pH adjusted to 4.7 with IN NaOH. 3.185 mg of l-Ethyl-3- (3- dimethylaminopropyl) carbodiimide hydrochloride (EDC) was added to the glucuronic solution and the solution was immediately transferred to the PEI solution by drop-wise addition. 4. The reaction pH was maintained at 4.7 by addition of0.1N NaOH for 3 hours.5. The preparation was dialyzed extensively against deionized H2O. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With sodium hydroxide; 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride In water for 3h; | 9 Example 9 Preparation of crlucopyranosyl- polyallylamine (GIu-PA)1. 100 mg of polyallylamine hydrochloride dissolved in 1 mL of deionized H2O. The pH was adjusted to 4.7 with IN NaOH.2. 184 mg of glucuronic acid dissolved in 1.2 ml of deionized H20and the pH adjusted to 4.7 with IN NaOH. 3. 180 mg of l-Ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) was added to the glucuronic solution and the solution was immediately transferred drop-wise to the PA solution.4. The reaction pH was maintained at 4.7 by addition of 0.1N NaOH for 3 hours.5. The preparation was dialyzed extensively against deionized H, 20V . |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride; sodium hydroxide; 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride In water for 3h; | 1.1.1 1. 5 g of Carboxymethylcellulose (CMC) sodium salt dissolved in 200 mL of deionized H2O. 2. 93 g of Ethylenediamine dihydrochloride (EDA) was added to the CMC solution and the pH adjusted to 4.7 with IN NaOH. 3. 6.7 g of l-Ethyl-3-(3- dimethylaminopropyl) carbodiimide hydrochloride (EDC) was added to the solution and the pH was maintained at 4.7 with IN HCl for 3 hours. 4. The preparation was dialyzed extensively against deionized H20 and lyophilized.5. 2 g of AE-CMC dissolved in 200 mL of deionized H2O. 6. 3 g of glucuronic acid dissolved in the AE-CMC solution. The pH adjusted with ION NaOH to 4.7.7. 6 g of EDC added to the solution and pH was maintained at 4.7 by addition of IN HCl. The reaction proceeded for 3 hours .8. The solution was dialyzed extensively against deionized H20 followed by lyophilization. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride; sodium hydroxide; 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride In water for 3h; | 1.1.10 1. 5 g of Dextran (MW 5x106 - 40x106) was dissolved in 140 mL of 1.35 M Chloroacetic acid 2. 27 mL ION NaOH and 10 mL of H2O were added and the reaction allowed to proceed overnight at 400C.3. The reaction was terminated by addition of 7.5 mL of 2 M NaH2PO4 and neutralized with 5N HCl. 4. The CM-Dextran was dialyzed extensively against deionized H20 and lyophilized.5. The lyophilized CM-Dextran was dissolved in 200 mL deionized H20.6. 93 g of EDA was added to the CM-Dextran solution and the pH adjusted to 4.7 with INNaOH.7. 6.7 g of EDC was added to the solution and the pH was maintained at 4.7 with IN HCl for 3 hours . 8. The preparation AECM-Dextran was dialyzed extensively against deionized H2O and lyophilized. 9. 2 g of AECM-Dextran was dissolved in 100 mL of deionized H2O.10. 3 g of glucuronic acid dissolved in the AECM- Dextran solution. The pH adjusted with ION NaOH to 4.7.11. 6 g of EDC added to the solution and pH was maintained at 4.7 by addition of IN HCl. The reaction proceeded for 3 hours .12. The preparation Glu-AECM-Dextran was dialyzed extensively against deionized H20 followed by lyophilization . |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride; sodium hydroxide; 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride In water for 3h; | 1.1.8 1. 5 g of Ficoll 400 was dissolved in 70 mL of 1.35 M Chloroacetic acid2. 13.5 mL ION NaOH and 5 mL of H2O were added and the reaction allowed to proceed overnight at 40° C.3. The reaction was terminated by addition of 3.75 mL of 2 M NaH2PO4 and neutralized with 5NHCl.4. The CM-Ficoll was dialyzed extensively against deionized H20 and lyophilized.5. The lyophilized CM-Ficoll was dissolved in dissolved in 200 mL deionized H2O.6. 93 g of EDA was added to the CM-Ficoll solution and the pH adjusted to 4.7 with IN NaOH.7. 6.7 g of EDC was added to the solution and the pH was maintained at 4.7 with IN HCl for 3 hours . 8. The preparation AECM-Picoll was dialyzed extensively against deionized H20 and lyophilized.9. 2 g of AECM-Ficoll was dissolved in 100 mL of deionized H20.10. 3 g of glucuronic acid dissolved in the AECM- Ficoll solution. The pH adjusted with ION NaOH to 4.7.11. 6 g of EDC added to the solution and pH was maintained at 4.7 by addition of IN HCl. The reaction proceeded for 3 hours .12. The preparation Glu-AECM-Ficoll was dialyzed extensively against deionized H2O followed by lyophilization. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride; sodium hydroxide; 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride In water for 3h; | 1.1.13 1. 5 g Gum Arabic dissolved in 100 mL of deionized H2O.2. 6.6 g of sodium periodate was added to the GA solution and pH adjusted to 5.0 with 2N NaOH.3. The reaction was covered with foil and stirred for 24 hours at 4° C. 4. The dialdehyde GA was dialyzed exhaustively against water. 5. 25 g of EDA was added to the solution followed by the addition of 2.5 g sodium cyanoborohydride dissolved in 10 mL of dimethylformamide and 15 mL of I N NaOH. The reaction mixture was stirred overnight at room temperature .6. After extensive dialysis the resulting aminoethylated GA (AE-GA) was lyophilized.7. 2 g of AE-GA dissolved in 60 mL deionized H2O followed by addition of 2 g glucuronic acid in20 mL deionized H2O. The pH adjusted with 2.5N NaOH to 4.7.8. 4 g of EDC was added to the solution and the pH was maintained at 4.7 with IN HCl for 3 hours .9. The GIu-AE-GA preparation was dialyzed against deionized H20 and lyophilized. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride; 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride In water for 3h; | 1.1.22 1. I g Gum Arabic dissolved in 20 mL of deionizedH2O. 2. 1 g of 1, 10-diaminodecane (DAD) dissolved in 6 mL of methanol .3. The DAD and GA solutions were mixed and pH adjusted to 4.7 using ION HCl.4. 1 g of EDC was added to the solution and the pH was maintained at 4.7 with IN HCl for 3 hours . 5. The DAD-GA preparation was dialyzed extensively against deionized H2O and lyophilized.6. The DAD-GA was dissolved in 100 mL of deionized H2O.7. 0.5 g of glucuronic acid was added to the DAD- GA solution and the pH adjusted to 4.7.8. 0.5 g of EDC added to the solution and pH was maintained at 4.7 by addition of IN HCl. The reaction proceeded for 3 hours .9. The solution was dialyzed extensively against deionized |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride; sodium hydroxide; 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride In water for 3h; | 1.1.11 1. 0.5 g DEAE-Dextran (purchased from Sigma-Aldrich) dissolved in 200 mL of deionized H2O.2. 0.3 g glucuronic acid added to the DEAE-Dex and pH adjusted to 4.7 with 2N NaOH.3. 0.6 g EDC added to the solution and pH was maintained at 4.7 for 3 hours with 0.1N HCl4. The Glu-DEAE-Dex solution was extensively dialyzed and lyophilized. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride; sodium hydroxide; 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride In water for 3h; | 1.1.19 1. 14 g CMC dissolved in 448 mL of deionized H2O and pH adjusted to 5.0.2. 112 mL of Trimethylolpropane tris [poly (propylene glycol) amine terminated] ether was diluted with an equal volume of H2O and the pH adjusted to 5.2 with ION HCl3. 14 g of EDC was used for coupling the reaction and the pH maintained at 4.7 by addition of IN HCl for 3 hours4. The TMA-CMC preparation was exhaustively dialyzed against deionized H2O followed by the reduction of volume to 400 mL by partial lyophilization.5. 1O g glucuronic acid was added to the TMA-CMC solution and pH adjusted to 5.0 with ION NaOH. 6 . 1O g EDC added to the solution and pH was maintained at 4.7 by addition of IN HCl. The reaction proceeded for 3 hours .7. The solution was dialyzed extensively against deionized H2O and lyophilized. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride; sodium hydroxide; 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride In water for 3h; | 1.1.23 1. 0.5 g GA dissolved in 8 mL of deionized H2O and pH adjusted to 5.2. 5 mL of Trimethylolpropane tris [poly (propylene glycol) amine terminated] ether was diluted with an equal volume of H2O and the pH adjusted to 5.2 with ION HCl3. 0.5 g of EDC was used for coupling the reaction and the pH maintained at 4.7 by addition of IN HCl for 3 hours4. The TMA-GA preparation was exhaustively dialyzed against deionized H2O followed by lyophilization.5. 210 mg glucuronic acid was added to 40 mL of TMA-GA solution and pH adjusted to 4.6 withION NaOH.6. 200 mg EDC added to the solution and pH was maintained at 4.7 by addition of IN HCl. The reaction proceeded for 3 hours . 7. The solution was dialyzed extensively against deionized H2O and lyophilized. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With bovine liver beta-glucuronidase; at 37℃;pH 5.0;Aqueous acetate buffer;Enzyme kinetics; | Thus, In order to optimize the enzyme cleavage kinetics of 1, the present invention provides the incorporation of a nitrophenyl self-immolative linker. The kinetics of previous galactosidase sensitive agents (EGad and EGadMe) (See, e.g., Moats et al., Chem., Int. Ed. Engl. 1997, 36, 726-728; Louie et al., Nat. Biotechnol. 2000, 18, 321-325) were impedingly slow, prompting discovery of the compositions and methods provided by the present invention. Thus, in some embodiments, the present invention provides an MR contrast agent that is modulated by changing q, the number of inner-sphere, Gd(III) coordinated water molecules. The use of a linker longer than the hydroxyethyl structure used in EGad (See, e.g., FIG. 3B) may preclude efficient water blockage by the sugar. However, the seven-coordinate DO3A analogs have reduced relaxivities due to coordination of endogenous bidentate anions such as carbonate (See, e.g., Bruce et al., J. Am. Chem. Soc. 2000, 122, 9674-9684; Dickins et al., J. Am. Chem. Soc. 2002, 124, 12697-12705; Messeri et al., Chem. Comm. 2001, 2742-2743; Supkowski et al., Inorg. Chem. 1999, 38, 5616-5619). Thus, the present invention provides the seven coordinate chelate structure of 1 that allows bidentate anion binding to occur. In some embodiments, upon enzymatic cleavage, the aminoethyl arm binds the metal center expelling the anion and generating an octadentate center with q=1, thus creating compound 2 (See, e.g., FIG. 3A). Octadentate complexes such as 2 bind anions with a much lower affinity (See, e.g., Supkowski et al., Inorg. Chem. 1999, 38, 5616-5619; Burai et al.,. Mag. Reson. Med. 1997, 38, 146-150) (3 orders of magnitude for carbonate). Thus, it is contemplated that, in some embodiments, water access should approach that of a q=1 complex. Therefore, in some embodiments, 2 has a higher relaxivity than the nominally q=0 1 in the presence of endogenous anions and the agent goes from low relaxivity (dark) to high relaxivity (bright) in the presence of beta-glucuronidase. The original investigations involving the beta-galactosidase activated agents, EGad, 4, and EGadMe, 5, (See, e.g., FIG. 3B) used an ?aqueous? synthetic route to obtain the desired complexes (See, e.g., Moats et al., Chem., Int. Ed. Engl. 1997, 36, 726-728; Louie et al., Nat. Biotechnol. 2000, 18, 321-325). In this scheme, the sugar/cyclen conjugate was deprotected in aqueous methanol and the acetate arms were added in alkaline water. The complex was isolated from this mixture using anion exchange. The large quantities of ammonium acetate used for ion exchange proved difficult to completely remove and hindered the subsequent metallation reaction. Furthermore, the one-pot approach did not permit comprehensive characterization of intermediates. For these reasons, an ?organic? synthetic route to the more complex compound 1 was employed (See, e.g., Schemes 2-6, below). This procedure permitted facile characterization with increased reproducibility. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
94.7% | In methanol at 20 - 55℃; | 9 Example 9 glucuronate [Show Image] 3.01 g base (10 mmoles) are dissolved in 20 ml methanol at 25°C. The mixture is heated at 55°C and a solution of 2.13 g D-glucuronic acid (11 mmoles) in 20 ml water is added over 10 min. The solution is seeded at 50°C and crystallization takes rapidly place. The slurry becomes rapidly too thick. The suspension is gradually diluted with 20 ml methanol / water = 1 / 1 during cooling and stirred over night at room temperature. The crystals are collected after filtration and washing with 20 ml methanol / water = 1 /1. Then a re-slurry wash with 20 ml acetone is performed. The solid is dried at 80°C and ca. 10 mbar for 20 Yield: 4.68 g white powder (94.7 %) Elemental analysis: Calc.: 53.32 % C; 6.71 % H; 14.13 % N; 25.83 % OFound: 53.17 % C; 6.61 % H; 14.05 % N; 26.01 % OWater assay: 0.19 % m/m |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
In isopropyl alcohol; at 40℃; for 1h; | Example 4; Preparation of Rasagiline Glucuronate Amorphous Form150 mg of <strong>[136236-51-6]rasagiline</strong> base was dissolved in 1 mL of 2-propanol. D-glucuronic acid (184 mg) was added and the mixture was stirred for 1 h at 40 C. The mixture was allowed to cool to ambient temperature and stirred for 24 hours at this temperature. The mixture was filtered and dried at under ambient conditions.Analytical data: XRD: Amorphous form, see FIG. 7. IR: see FIG. 8. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With sulfuric acid; water at 100℃; for 1h; | 1.a To 100 g of Okinawa Nemacystus decipiens, 1000 ml of distilled water was added, and extraction was conducted at 100° C. for 1 hour. The obtained extract was cooled, and then filtered by suction, electrodialyzed (desalinated), and lyophilized to obtain 2 g of fucoidan fractions. This fucoidan was hydrolyzed with an aqueous solution containing 2NH2SO4 at 100° C. for 1 hour. The obtained aqueous solution was neutralized by 2 N NaOH, and fluorescently labeled by ABEE to prepare a monosaccharide analysis sample. It was confirmed that the composition of the constituent sugars was sulfated fucose:glucuronic acid:fucose:xylose=49.3:4.9:12.1:1 (FIG. 1).Column: Cosmosil C18 AR-II (4.6 mm×250 mm) Mobile phase: 0.2 M potassium borate buffer containing 10% acetonitrile Flow rate: 1.0 ml/min. Temperature: 45° C.Detection: fluorescence detector (Shimadzu Corporation), Ex: 305 nm, Em: 360 nm |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride In 1,4-dioxane; water at 80℃; for 3h; | 3.4. Acid hydrolysis and sugar identification Each saponin (3 mg) was heated in 3 mL of 10% 1:1 HCl-dioxane at 80 °C for 3 h. The solvent was removed in vacuo, and the residue was partitioned between EtOAc and H2O to give aglycone and sugar, respectively. The sugar components in the aqueous layer were analyzed by silica gel TLC by comparison with standard sugars. The solvent system was 2:1:0.2 CH2Cl2-MeOH-H2O, and spots were visualized by spraying with 9:0.5:0.5 95% EtOH-H2SO4-anisaldehyde, then heated at 105 °C for 5 min. For sugars of the saponins, the Rf values of glucuronic acid, glucose, galactose, and arabinose by TLC were 0.15, 0.30, 0.32, and 0.5, respectively. The results were confirmed by GC analysis as follows. The aqueous layer was evaporated to dryness to give a residue that was dissolved in anhydrous pyridine (100 μL) and then mixed with a pyridine solution of 0.1 M l-cysteine methyl ester hydrochloride (100 μL). After warming at 60 °C for 2 h, trimethylsilylimidazole solution was added and warmed at 60 °C for 2 h. The mixture was then evaporated in vacuo to give a dried product, which was partitioned between n-hexane and H2O. The n-hexane layer was filtered and analyzed by GC. The absolute configurations of the monosaccharides were confirmed to be l-arabinose, d-glucuronic acid, d-glucose, and d-galactose by comparison of the retention times of persilylated monosaccharide derivatives with those of standard samples. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride In 1,4-dioxane; water at 80℃; for 3h; | 3.4. Acid hydrolysis and sugar identification Each saponin (3 mg) was heated in 3 mL of 10% 1:1 HCl-dioxane at 80 °C for 3 h. The solvent was removed in vacuo, and the residue was partitioned between EtOAc and H2O to give aglycone and sugar, respectively. The sugar components in the aqueous layer were analyzed by silica gel TLC by comparison with standard sugars. The solvent system was 2:1:0.2 CH2Cl2-MeOH-H2O, and spots were visualized by spraying with 9:0.5:0.5 95% EtOH-H2SO4-anisaldehyde, then heated at 105 °C for 5 min. For sugars of the saponins, the Rf values of glucuronic acid, glucose, galactose, and arabinose by TLC were 0.15, 0.30, 0.32, and 0.5, respectively. The results were confirmed by GC analysis as follows. The aqueous layer was evaporated to dryness to give a residue that was dissolved in anhydrous pyridine (100 μL) and then mixed with a pyridine solution of 0.1 M l-cysteine methyl ester hydrochloride (100 μL). After warming at 60 °C for 2 h, trimethylsilylimidazole solution was added and warmed at 60 °C for 2 h. The mixture was then evaporated in vacuo to give a dried product, which was partitioned between n-hexane and H2O. The n-hexane layer was filtered and analyzed by GC. The absolute configurations of the monosaccharides were confirmed to be l-arabinose, d-glucuronic acid, d-glucose, and d-galactose by comparison of the retention times of persilylated monosaccharide derivatives with those of standard samples. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With oxygen for 0.5h; | ||
75 %Chromat. | With oxygen In water at 70℃; for 4h; Autoclave; High pressure; | Oxidation reactions General procedure: The reactions were carried out in 75mL stainless steel reactors (Series 5000 Multiple Reactor System, Parr Instrument Co., USA). The test reactor unit is a magnetically stirred high-pressure reactor equipped with a Parr 4843 controller for the setup and control of reaction temperature and stirring speed. Reactor pressure measurements were accomplished via a pressure transducer attached to the reactor. Temperature, pressure and stirring speed are recorded by a Space View data acquisition system. The total volume of carbohydrate aqueous solution used was 10mL and the charged weight of catalyst was about 20mg. The catalytic reactions were carried out for the desired time or temperature without any base addition. The oxygen pressure was of 15bar in all experiments |
With glucose oxidase In aq. phosphate buffer at 35℃; for 0.5h; Enzymatic reaction; |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With trifluoroacetic acid at 120℃; for 3h; | 2.2 Extraction, isolation and purification of polysaccharide The polysaccharide was hydrolyzed by 2M TFA at 120°C for 3h into monosaccharides under airtight condition, and the monosaccharides were conventionally converted into the alditol acetates as described (Xin et al., 2012) and were analyzed by gas chromatography (GC, Agilent 6890, USA) equipped with a HP-5 column (30m×0.25mm×0.25μm) and flame-ionization detector (FID). The operation was performed using the following conditions: 160°C for 2min, then to 200°C at 6°C/min, then to 215°C at 0.2°C/min, then to 240°C at 6°C/min for 30min. Nitrogen was used as the carrier gas at 1mL/min; injection temperature was 250°C; detector temperature was 300°C. Monosaccharides identification was done by comparison with reference monosaccharides. The relative molar proportions were calculated by the area normalization method. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With b-glucuronidase In aq. buffer at 37℃; for 5h; Enzymatic reaction; | Hydrolysis of compounds 1 and 2 Compound 2 (5 mg) was dissolved in0.5 ml of KH2PO4/KOH buffer solution(pH 4.90) before 0.2 ml of b-glucuronidasesolution was added, and incubated at378C for 5 h. The reaction mixture (about0.7 ml) was mixed with 3ml of methanol,vortexed at 1500 rpm for 5min, andcentrifuged at 10,000 rpm for 10 min. Thesupernatant was transferred to a clean testtube and dried under a gentle flow ofnitrogen gas at 40 8C. The residue waspartitioned between EtOAc and H2O(4 £ 1 ml). The water-soluble part wasreduced to dryness and then analyzed byco-TLC over silica gel (EtOAc-EtOH-H2O-HCOOH 6:4:1:1), by comparisonwith authentic sugar (D-glucuronose), andthe EtOAc-soluble part was analyzed byLC-MSn compared with compound 2[Agilent 1100 series HPLC systemcoupled with Finnigan LCQ Advantageion trap mass spectrometer via ESI interface,Grace Allsphere ODS-2 column(5 mm, 4.6mm £ 250 mm), mobile phaseconsisted of acetonitrile (A) and watercontaining 0.1% (v/v) formic acid (B) inthe following gradient program: 0 min,10% A; 60 min, 60% A; flow rate 1 ml/min], which confirmed the presence ofD-glucuronic acid.Compound 1 (2 mg) and compound 2(the dried EtOAc extract) were hydrolyzedwith 1 N HCl (2 ml) for 6 h at 1008C,respectively. After lyopilization, the dryresidue was partitioned between Et2O andH2O (3 £ 2 ml). The dried water extractand 3 mg of L-cysteine methyl ester hydrochloride were dissolved in 0.3 ml ofanhydrous pyridine, stirred at 60 8C for 2 h.The reaction mixture was dried undervacuum and was trimethylsilylated with0.5 ml of HMDS-TMCS-pyridine (3:1:9)at 60 8C for 2 h. The mixture was thenconcentrated and partitioned betweenn-hexane and H2O. The n-hexane extractwas analyzed by GC under the followingconditions: Varian CP-3800 GC, DB-5 GCcolumn (30m £ 0.25mm £ 0.25mm), columntemperature increased from 50 to260 8C at 68C/min, carrier gas N2. The tRvalues (min) of trimethylsilyl ether derivativesof authentic sugars (L-Ara, D-Ara,and L-Rha) prepared in a similar way were24.12, 24.51 and 24.96 min, respectively.L-Ara and L-Rha were detected fromcompounds 1 (tR 24.13 min) and 2 (tR24.92 min). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With trifluoroacetic acid In water at 95℃; for 3h; | Acid hydrolysis and GC analysis General procedure: Each compound (3 mg) was hydrolyzed with 2 N aq. CF3COOH (5 ml) for 3 h at 95 °C. After extraction with CH2Cl2 (3 × 5 ml), the aq. layer was repeatly evaporated to dryness with MeOH until neutral, and then analysed by TLC over silica gel (CHCl3-MeOH-H2O 8/5/1) by comparaison with authentic samples. Futhermore, the residue of sugars was dissolved in anhydrous pyridine (100 μl), and l-cysteine methyl ester hydrochloride (0.06 mol/l) was added. The mixture was stirred at 60 °C for 1 h, then 150 μl of HMDS-TMCS (hexamethyldisilazane-trimethylchlorosilane 3:1) was added, and the mixture was stirred at 60 °C for another 30 min. The precipitate was centrifuged off, and the supernatant was concentrated under N2 stream. The residue was partioned between n-hexane and H2O (0.1 ml each), and the hexane layer (1 μl) was analysed by GC (Hara et al., 1987). The absolute configurations were determined by comparing the retention times with thiazolidine derivatives prepared in a similar way from standard sugars (Sigma-Aldrich): d-glucuronic acid, d-xylose, d-galactose, d-glucose, d-fucose, l-rhamnose for 1, 2; d-glucuronic acid, d-xylose, d-galactose, d-fucose, d-quinovose, l-rhamnose for 3 and d-galactose, d-glucose for 4 were characterized by co-injection of the hydrosylate with standard silylated samples having tR 15.2 min (d-glucuronic acid), 13.4 min (d-xylose), 19.6 min (d-galactose), 18.6 min (d-glucose), 12.1 min (d-fucose), 13.1 min (l-rhamnose) and 11.1 min (d-quinovose). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With trifluoroacetic acid In water at 95℃; for 3h; | Acid hydrolysis and GC analysis General procedure: Each compound (3 mg) was hydrolyzed with 2 N aq. CF3COOH (5 ml) for 3 h at 95 °C. After extraction with CH2Cl2 (3 × 5 ml), the aq. layer was repeatly evaporated to dryness with MeOH until neutral, and then analysed by TLC over silica gel (CHCl3-MeOH-H2O 8/5/1) by comparaison with authentic samples. Futhermore, the residue of sugars was dissolved in anhydrous pyridine (100 μl), and l-cysteine methyl ester hydrochloride (0.06 mol/l) was added. The mixture was stirred at 60 °C for 1 h, then 150 μl of HMDS-TMCS (hexamethyldisilazane-trimethylchlorosilane 3:1) was added, and the mixture was stirred at 60 °C for another 30 min. The precipitate was centrifuged off, and the supernatant was concentrated under N2 stream. The residue was partioned between n-hexane and H2O (0.1 ml each), and the hexane layer (1 μl) was analysed by GC (Hara et al., 1987). The absolute configurations were determined by comparing the retention times with thiazolidine derivatives prepared in a similar way from standard sugars (Sigma-Aldrich): d-glucuronic acid, d-xylose, d-galactose, d-glucose, d-fucose, l-rhamnose for 1, 2; d-glucuronic acid, d-xylose, d-galactose, d-fucose, d-quinovose, l-rhamnose for 3 and d-galactose, d-glucose for 4 were characterized by co-injection of the hydrosylate with standard silylated samples having tR 15.2 min (d-glucuronic acid), 13.4 min (d-xylose), 19.6 min (d-galactose), 18.6 min (d-glucose), 12.1 min (d-fucose), 13.1 min (l-rhamnose) and 11.1 min (d-quinovose). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With trifluoroacetic acid In water at 95℃; for 3h; | Acid hydrolysis and GC analysis General procedure: Each compound (3 mg) was hydrolyzed with 2 N aq. CF3COOH (5 ml) for 3 h at 95 °C. After extraction with CH2Cl2 (3 × 5 ml), the aq. layer was repeatly evaporated to dryness with MeOH until neutral, and then analysed by TLC over silica gel (CHCl3-MeOH-H2O 8/5/1) by comparaison with authentic samples. Futhermore, the residue of sugars was dissolved in anhydrous pyridine (100 μl), and l-cysteine methyl ester hydrochloride (0.06 mol/l) was added. The mixture was stirred at 60 °C for 1 h, then 150 μl of HMDS-TMCS (hexamethyldisilazane-trimethylchlorosilane 3:1) was added, and the mixture was stirred at 60 °C for another 30 min. The precipitate was centrifuged off, and the supernatant was concentrated under N2 stream. The residue was partioned between n-hexane and H2O (0.1 ml each), and the hexane layer (1 μl) was analysed by GC (Hara et al., 1987). The absolute configurations were determined by comparing the retention times with thiazolidine derivatives prepared in a similar way from standard sugars (Sigma-Aldrich): d-glucuronic acid, d-xylose, d-galactose, d-glucose, d-fucose, l-rhamnose for 1, 2; d-glucuronic acid, d-xylose, d-galactose, d-fucose, d-quinovose, l-rhamnose for 3 and d-galactose, d-glucose for 4 were characterized by co-injection of the hydrosylate with standard silylated samples having tR 15.2 min (d-glucuronic acid), 13.4 min (d-xylose), 19.6 min (d-galactose), 18.6 min (d-glucose), 12.1 min (d-fucose), 13.1 min (l-rhamnose) and 11.1 min (d-quinovose). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With trifluoroacetic acid In water at 95℃; for 3h; | Acid hydrolysis and GC analysis General procedure: Each compound (3 mg) was hydrolyzed with 2 N aq. CF3COOH (5 ml) for 3 h at 95 °C. After extraction with CH2Cl2 (3 × 5 ml), the aq. layer was repeatly evaporated to dryness with MeOH until neutral, and then analysed by TLC over silica gel (CHCl3-MeOH-H2O 8/5/1) by comparaison with authentic samples. Futhermore, the residue of sugars was dissolved in anhydrous pyridine (100 μl), and l-cysteine methyl ester hydrochloride (0.06 mol/l) was added. The mixture was stirred at 60 °C for 1 h, then 150 μl of HMDS-TMCS (hexamethyldisilazane-trimethylchlorosilane 3:1) was added, and the mixture was stirred at 60 °C for another 30 min. The precipitate was centrifuged off, and the supernatant was concentrated under N2 stream. The residue was partioned between n-hexane and H2O (0.1 ml each), and the hexane layer (1 μl) was analysed by GC (Hara et al., 1987). The absolute configurations were determined by comparing the retention times with thiazolidine derivatives prepared in a similar way from standard sugars (Sigma-Aldrich): d-glucuronic acid, d-xylose, d-galactose, d-glucose, d-fucose, l-rhamnose for 1, 2; d-glucuronic acid, d-xylose, d-galactose, d-fucose, d-quinovose, l-rhamnose for 3 and d-galactose, d-glucose for 4 were characterized by co-injection of the hydrosylate with standard silylated samples having tR 15.2 min (d-glucuronic acid), 13.4 min (d-xylose), 19.6 min (d-galactose), 18.6 min (d-glucose), 12.1 min (d-fucose), 13.1 min (l-rhamnose) and 11.1 min (d-quinovose). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
34.6% | With phosphovanadomolybdic acid; oxygen In water at 150℃; for 3h; | General procedure: The oxidative conversions of cellulose by various HPA catalysts were carried out in a 75 mL Teflon-lined stainless autoclave at 453 K for 3 h under 0.4-2 MPa O2 with a stirring rate of 600 rpm. Typically, the reaction mixture comprised 20 mL of H2O, 0.2 g of α-cellulose powder (containing 1.23 mmol glucose units), and 0.1 mmol of HPA catalyst. In the reactions with other substrates, a fixed amount of reactant (200 mg) and the typical reaction conditions were used unless otherwise specified. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
72.6% | With tetrabutyl ammonium fluoride In tetrahydrofuran; N,N-dimethyl-formamide at 0 - 20℃; for 18h; Inert atmosphere; | Synthesis of diclofenac 1-β acyl glucuronide. Benzyl bromide (5.15 mL, 43.27 mmol) was added dropwise over 5 min, with stirring, to a solutionof D-glucuronic acid 1 (8 g, 41.21 mmol) in DMF (50 mL) and 1 M TBAF in THF (45.3 mL, 45.33mmol) cooled to 0°C under nitrogen. The resulting solution was left for 18 h, during which time itwas allowed to warm to ambient temperature.The solvents were firstly evaporated and then co-evaporated with toluene (100 mL × 4) to give anorange oil containing residual DMF. The oil was triturated with toluene (200 mL), the solventdecanted, and the crude product was purified by flash-column silica chromatography by loading itonto the column with DCM and performing gradient elution with 0-10% MeOH in EtOAc. Purefractions were evaporated to dryness to afford benzyl , -D-glucuronate 2 (8.5 g, 72.6%) as acolourless gum. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With sulfuric acid In 1,4-dioxane; water at 90℃; for 3h; | 3.4 Acid hydrolysis and monosaccharide composition of floraassamsaponins I-VIII General procedure: To determine the absolute configurations of constituent monosaccharides of 1-8, the reported method10 was used with slight modifications as follows. Compounds 1-8 (1.0-2.0 mg) were dissolved in 5% aqueous H2SO4/1,4-dioxane (1:1, v/v, 2.0 mL), and each solution was heated at 90°C for 3h. After extraction three times with EtOAc, the aqueous layer was neutralized with Amberlite IRA-400 (OH- form). After drying in vacuo, the residue was dissolved in pyridine (0.1mL) containing l-cysteine methyl ester hydrochloride (0.5mg) and heated at 60°C for 1h. A solution of o-tolylisothiocyanate (0.5mg) in pyridine (0.1mL) was added to the mixture and heated at 60°C for 1h. The reaction mixture was analyzed by reversed-phase HPLC [column: COSMOSIL 5C18-AR-II (Nacalai Tesque), 250×4.6mm i.d. (5μm); mobile phase: MeCN/0.05M H3PO4 (18:82, v/v); detection: UV (250nm); flow rate: 0.8mL/min; column temperature: 35°C] to identify the derivatives of constituent monosaccharides (d-galactose, d-glucose, d-glucuronic acid, l-arabinose, and l-rhamnose) in 1-8 by comparison of their retention times with those of authentic samples (tR: d-galactose; 44.1min, l-galactose; 46.9min, d-glucose; 51.6min, l-glucose; 50.0min, d-glucuronic acid; 52.7min, l-glucuronic acid; 44.0min, d-arabinose; 63.4min, l-arabinose; 58.4min, d-rhamnose; 88.4min, l-rhamnose; 92.7min). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With sulfuric acid In 1,4-dioxane; water at 90℃; for 3h; | 3.4 Acid hydrolysis and monosaccharide composition of floraassamsaponins I-VIII General procedure: To determine the absolute configurations of constituent monosaccharides of 1-8, the reported method10 was used with slight modifications as follows. Compounds 1-8 (1.0-2.0 mg) were dissolved in 5% aqueous H2SO4/1,4-dioxane (1:1, v/v, 2.0 mL), and each solution was heated at 90°C for 3h. After extraction three times with EtOAc, the aqueous layer was neutralized with Amberlite IRA-400 (OH- form). After drying in vacuo, the residue was dissolved in pyridine (0.1mL) containing l-cysteine methyl ester hydrochloride (0.5mg) and heated at 60°C for 1h. A solution of o-tolylisothiocyanate (0.5mg) in pyridine (0.1mL) was added to the mixture and heated at 60°C for 1h. The reaction mixture was analyzed by reversed-phase HPLC [column: COSMOSIL 5C18-AR-II (Nacalai Tesque), 250×4.6mm i.d. (5μm); mobile phase: MeCN/0.05M H3PO4 (18:82, v/v); detection: UV (250nm); flow rate: 0.8mL/min; column temperature: 35°C] to identify the derivatives of constituent monosaccharides (d-galactose, d-glucose, d-glucuronic acid, l-arabinose, and l-rhamnose) in 1-8 by comparison of their retention times with those of authentic samples (tR: d-galactose; 44.1min, l-galactose; 46.9min, d-glucose; 51.6min, l-glucose; 50.0min, d-glucuronic acid; 52.7min, l-glucuronic acid; 44.0min, d-arabinose; 63.4min, l-arabinose; 58.4min, d-rhamnose; 88.4min, l-rhamnose; 92.7min). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With sulfuric acid In 1,4-dioxane; water at 90℃; for 3h; | 3.4 Acid hydrolysis and monosaccharide composition of floraassamsaponins I-VIII General procedure: To determine the absolute configurations of constituent monosaccharides of 1-8, the reported method10 was used with slight modifications as follows. Compounds 1-8 (1.0-2.0 mg) were dissolved in 5% aqueous H2SO4/1,4-dioxane (1:1, v/v, 2.0 mL), and each solution was heated at 90°C for 3h. After extraction three times with EtOAc, the aqueous layer was neutralized with Amberlite IRA-400 (OH- form). After drying in vacuo, the residue was dissolved in pyridine (0.1mL) containing l-cysteine methyl ester hydrochloride (0.5mg) and heated at 60°C for 1h. A solution of o-tolylisothiocyanate (0.5mg) in pyridine (0.1mL) was added to the mixture and heated at 60°C for 1h. The reaction mixture was analyzed by reversed-phase HPLC [column: COSMOSIL 5C18-AR-II (Nacalai Tesque), 250×4.6mm i.d. (5μm); mobile phase: MeCN/0.05M H3PO4 (18:82, v/v); detection: UV (250nm); flow rate: 0.8mL/min; column temperature: 35°C] to identify the derivatives of constituent monosaccharides (d-galactose, d-glucose, d-glucuronic acid, l-arabinose, and l-rhamnose) in 1-8 by comparison of their retention times with those of authentic samples (tR: d-galactose; 44.1min, l-galactose; 46.9min, d-glucose; 51.6min, l-glucose; 50.0min, d-glucuronic acid; 52.7min, l-glucuronic acid; 44.0min, d-arabinose; 63.4min, l-arabinose; 58.4min, d-rhamnose; 88.4min, l-rhamnose; 92.7min). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With sulfuric acid In 1,4-dioxane; water at 90℃; for 3h; | 3.4 Acid hydrolysis and monosaccharide composition of floraassamsaponins I-VIII General procedure: To determine the absolute configurations of constituent monosaccharides of 1-8, the reported method10 was used with slight modifications as follows. Compounds 1-8 (1.0-2.0 mg) were dissolved in 5% aqueous H2SO4/1,4-dioxane (1:1, v/v, 2.0 mL), and each solution was heated at 90°C for 3h. After extraction three times with EtOAc, the aqueous layer was neutralized with Amberlite IRA-400 (OH- form). After drying in vacuo, the residue was dissolved in pyridine (0.1mL) containing l-cysteine methyl ester hydrochloride (0.5mg) and heated at 60°C for 1h. A solution of o-tolylisothiocyanate (0.5mg) in pyridine (0.1mL) was added to the mixture and heated at 60°C for 1h. The reaction mixture was analyzed by reversed-phase HPLC [column: COSMOSIL 5C18-AR-II (Nacalai Tesque), 250×4.6mm i.d. (5μm); mobile phase: MeCN/0.05M H3PO4 (18:82, v/v); detection: UV (250nm); flow rate: 0.8mL/min; column temperature: 35°C] to identify the derivatives of constituent monosaccharides (d-galactose, d-glucose, d-glucuronic acid, l-arabinose, and l-rhamnose) in 1-8 by comparison of their retention times with those of authentic samples (tR: d-galactose; 44.1min, l-galactose; 46.9min, d-glucose; 51.6min, l-glucose; 50.0min, d-glucuronic acid; 52.7min, l-glucuronic acid; 44.0min, d-arabinose; 63.4min, l-arabinose; 58.4min, d-rhamnose; 88.4min, l-rhamnose; 92.7min). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With sulfuric acid In 1,4-dioxane; water at 90℃; for 3h; | 3.4 Acid hydrolysis and monosaccharide composition of floraassamsaponins I-VIII General procedure: To determine the absolute configurations of constituent monosaccharides of 1-8, the reported method10 was used with slight modifications as follows. Compounds 1-8 (1.0-2.0 mg) were dissolved in 5% aqueous H2SO4/1,4-dioxane (1:1, v/v, 2.0 mL), and each solution was heated at 90°C for 3h. After extraction three times with EtOAc, the aqueous layer was neutralized with Amberlite IRA-400 (OH- form). After drying in vacuo, the residue was dissolved in pyridine (0.1mL) containing l-cysteine methyl ester hydrochloride (0.5mg) and heated at 60°C for 1h. A solution of o-tolylisothiocyanate (0.5mg) in pyridine (0.1mL) was added to the mixture and heated at 60°C for 1h. The reaction mixture was analyzed by reversed-phase HPLC [column: COSMOSIL 5C18-AR-II (Nacalai Tesque), 250×4.6mm i.d. (5μm); mobile phase: MeCN/0.05M H3PO4 (18:82, v/v); detection: UV (250nm); flow rate: 0.8mL/min; column temperature: 35°C] to identify the derivatives of constituent monosaccharides (d-galactose, d-glucose, d-glucuronic acid, l-arabinose, and l-rhamnose) in 1-8 by comparison of their retention times with those of authentic samples (tR: d-galactose; 44.1min, l-galactose; 46.9min, d-glucose; 51.6min, l-glucose; 50.0min, d-glucuronic acid; 52.7min, l-glucuronic acid; 44.0min, d-arabinose; 63.4min, l-arabinose; 58.4min, d-rhamnose; 88.4min, l-rhamnose; 92.7min). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With sulfuric acid In 1,4-dioxane; water at 90℃; for 3h; | 3.4 Acid hydrolysis and monosaccharide composition of floraassamsaponins I-VIII General procedure: To determine the absolute configurations of constituent monosaccharides of 1-8, the reported method10 was used with slight modifications as follows. Compounds 1-8 (1.0-2.0 mg) were dissolved in 5% aqueous H2SO4/1,4-dioxane (1:1, v/v, 2.0 mL), and each solution was heated at 90°C for 3h. After extraction three times with EtOAc, the aqueous layer was neutralized with Amberlite IRA-400 (OH- form). After drying in vacuo, the residue was dissolved in pyridine (0.1mL) containing l-cysteine methyl ester hydrochloride (0.5mg) and heated at 60°C for 1h. A solution of o-tolylisothiocyanate (0.5mg) in pyridine (0.1mL) was added to the mixture and heated at 60°C for 1h. The reaction mixture was analyzed by reversed-phase HPLC [column: COSMOSIL 5C18-AR-II (Nacalai Tesque), 250×4.6mm i.d. (5μm); mobile phase: MeCN/0.05M H3PO4 (18:82, v/v); detection: UV (250nm); flow rate: 0.8mL/min; column temperature: 35°C] to identify the derivatives of constituent monosaccharides (d-galactose, d-glucose, d-glucuronic acid, l-arabinose, and l-rhamnose) in 1-8 by comparison of their retention times with those of authentic samples (tR: d-galactose; 44.1min, l-galactose; 46.9min, d-glucose; 51.6min, l-glucose; 50.0min, d-glucuronic acid; 52.7min, l-glucuronic acid; 44.0min, d-arabinose; 63.4min, l-arabinose; 58.4min, d-rhamnose; 88.4min, l-rhamnose; 92.7min). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With acetic acid In water at 100℃; for 2h; | Acid hydrolysis Each saponin (2mg) was refluxed with 2N aq. CH3COOH (5ml) for 2h at 100°C. After extraction with CH3Cl (3× 5ml), the aqueous layer was repeatedly evaporated to dryness with MeOH until neutral, and then analyzed by TLC over silica gel (MeCOEt-isoPrOH-Me2CO-H2O 20:10:7:6) by comparison with authentic samples (l-rhamnose Rf 0.65; d-glucose Rf 0.40; d-glucuronic acid Rf 0.05) (Haddad et al., 2013; Voutquenne-Nazabadioko et al., 2013). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With trifluoroacetic acid In 1,4-dioxane at 105℃; for 2h; | Protocol for Acid Hydrolysis of Compound 3 Compound 3(2.0 mg) was dissolved in 4 ml of a mixture 1:1 of dioxane/TFA 2N and heated at105°C for 2 h. The solvent was evaporated and the remaining solution was extracted three times with 1 ml of CHCl3. TLC analysis of the organic phase revealed decomposition of the aglycone. The aqueous phase was dried and the residue re-dissolved in anhydrous pyridine. The sugars were derivatized with L-cysteine methyl ester hydrochloride (200 ml, 60°C, 1 h) and subsequently silylated with hexamethyldisilzane and chlorotrimethylsilane (Fluka) in pyridine (2:1:10, 300 ml; 60°C, 30 min). GC analysis on a capillary DB-225MScolumn (30 m x 0,25 mm i.d., 0,25 m; Agilent; column temp. 150°C for 2 min,then 5°C/min. to 210°C, then 10°C/min to 240°C). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride In methanol at 92℃; for 3h; | 3.4 Acid hydrolysis of compounds 1-3 General procedure: The experiment was implemented using the method previously described (Li et al., 2009). Compounds 1-3 (2.0mg) were treated with 3% HCl in MeOH (5mL) at 92°C for 3h, respectively. HCl and MeOH were removed under reduced pressure at 60°C. 5mL CHCl3/H2O (1:1) were used for extraction. The aqueous layers were evaporated to dryness and analyzed by TLC over silica gel (CHCl3-MeOH-H2O-HCOOH 15:5:1:1) by comparison with authentic samples (l-arabinose Rf 0.75; l-rhamnose Rf 0.65; d-xylose Rf 0.69; d-glucose Rf 0.55; d-glucuronic acid Rf 0.10). The results further confirmed that the structure of compounds 1-3. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride In methanol at 92℃; for 3h; | 3.4 Acid hydrolysis of compounds 1-3 General procedure: The experiment was implemented using the method previously described (Li et al., 2009). Compounds 1-3 (2.0mg) were treated with 3% HCl in MeOH (5mL) at 92°C for 3h, respectively. HCl and MeOH were removed under reduced pressure at 60°C. 5mL CHCl3/H2O (1:1) were used for extraction. The aqueous layers were evaporated to dryness and analyzed by TLC over silica gel (CHCl3-MeOH-H2O-HCOOH 15:5:1:1) by comparison with authentic samples (l-arabinose Rf 0.75; l-rhamnose Rf 0.65; d-xylose Rf 0.69; d-glucose Rf 0.55; d-glucuronic acid Rf 0.10). The results further confirmed that the structure of compounds 1-3. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride In water at 80℃; for 4h; | Acid hydrolysis and sugar analysisof compounds 1 and 2 General procedure: Each saponin (2 mg) was dissolved in 2NHCl (2 ml) and stirred at 808C for 4 h. Thereaction mixture was extracted withchloroform, and the aqueous layer wasevaporated to give a mixture of monosaccharides.The residue was dissolved inanhydrous pyridine (1 ml) followed by theaddition of 2mg of L-cysteine methyl esterhydrochloride (99%, Tokyo ChemicalIndustry Co., Ltd, Tokyo, Japan). Afterheating at 608C for 2 h, the solvent was eliminated under N2, and 0.2 ml trimethylsilylimidazole(Tokyo Chemical IndustryCo., Ltd, 99%) was added. Then themixture was heated at 608C for another 2 hand partitioned between n-hexane andwater. The organic layer was investigatedby GC under the following conditions:capillary column, HP-5 (30m £ 0.25mm£ 0.25 mm, Dikma Technologies, Beijing,China); FID detector with a temperature of2808C; injection temperature 2508C; initialtemperature 1608C, then raised to 2808C at58C/min, final temperature maintainedfor 10 min; carrier gas, N2. The standardsugars experienced the same reaction andGC conditions. The retention times of Dglucuronicacid, D-xylose, and L-arabinosewere 19.770, 32.257, and 33.284min,respectively. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride In water at 80℃; for 4h; | Acid hydrolysis and sugar analysisof compounds 1 and 2 General procedure: Each saponin (2 mg) was dissolved in 2NHCl (2 ml) and stirred at 808C for 4 h. Thereaction mixture was extracted withchloroform, and the aqueous layer wasevaporated to give a mixture of monosaccharides.The residue was dissolved inanhydrous pyridine (1 ml) followed by theaddition of 2mg of L-cysteine methyl esterhydrochloride (99%, Tokyo ChemicalIndustry Co., Ltd, Tokyo, Japan). Afterheating at 608C for 2 h, the solvent was eliminated under N2, and 0.2 ml trimethylsilylimidazole(Tokyo Chemical IndustryCo., Ltd, 99%) was added. Then themixture was heated at 608C for another 2 hand partitioned between n-hexane andwater. The organic layer was investigatedby GC under the following conditions:capillary column, HP-5 (30m £ 0.25mm£ 0.25 mm, Dikma Technologies, Beijing,China); FID detector with a temperature of2808C; injection temperature 2508C; initialtemperature 1608C, then raised to 2808C at58C/min, final temperature maintainedfor 10 min; carrier gas, N2. The standardsugars experienced the same reaction andGC conditions. The retention times of Dglucuronicacid, D-xylose, and L-arabinosewere 19.770, 32.257, and 33.284min,respectively. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With trifluoroacetic acid; In water; for 1h;Reflux; | General procedure: Individual solutions of 1-10 (each 5mg) and 11-12 (each 3mg) in 1M HCl (dioxane-H2O, 1:1, 10mL) were separately heated until reflux occurred, this being maintained for 1h. After removal of dioxane by evaporation, the solution was extracted with EtOAc (10mL×3). The EtOAc fractions from compounds 1, 2, and 3 were then separated by preparative NP-HPLC with CHCl3-MeOH-H2O (60:29:6, v/v/v) as eluent to afford 3-O-beta-D-glucuronopyranosyl-quillaic acid (21). EtOAc fractions from compounds 4-9 were separated by preparative NP-HPLC with CHCl3-MeOH (95:5, v/v) as eluent to afford 3beta,16alpha-dihydroxy-olean-12-en-23,28-dioic acid (18). The EtOAc fraction from compound 10 was separated by preparative NP-HPLC with CHCl3-MeOH (96:4, v/v) as the eluent to afford 3beta-hydroxy-16-oxo-28-nor-olean-12-en-23-oic acid (10a). EtOAc fractions from compounds 11-12 were separated by preparative NP-HPLC with CHCl3-MeOH (95:5, v/v) as eluent to afford 3,16alpha-dihydroxy-3,4-seco-olean-4(24),12-dien-23,28-dioic acid (20). Acid hydrolysis of compounds 1-3 (each 5mg) in 2M aqueous TFA using the same procedure afforded quillaic acid. Spectroscopic data including NMR and MS for all of the compounds except 10a were identical to those for authentic samples. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With oxygen In water at 70℃; for 4h; Autoclave; High pressure; | Oxidation reactions General procedure: The reactions were carried out in 75mL stainless steel reactors (Series 5000 Multiple Reactor System, Parr Instrument Co., USA). The test reactor unit is a magnetically stirred high-pressure reactor equipped with a Parr 4843 controller for the setup and control of reaction temperature and stirring speed. Reactor pressure measurements were accomplished via a pressure transducer attached to the reactor. Temperature, pressure and stirring speed are recorded by a Space View data acquisition system. The total volume of carbohydrate aqueous solution used was 10mL and the charged weight of catalyst was about 20mg. The catalytic reactions were carried out for the desired time or temperature without any base addition. The oxygen pressure was of 15bar in all experiments |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With oxygen In water at 70℃; for 4h; Autoclave; High pressure; | Oxidation reactions General procedure: The reactions were carried out in 75mL stainless steel reactors (Series 5000 Multiple Reactor System, Parr Instrument Co., USA). The test reactor unit is a magnetically stirred high-pressure reactor equipped with a Parr 4843 controller for the setup and control of reaction temperature and stirring speed. Reactor pressure measurements were accomplished via a pressure transducer attached to the reactor. Temperature, pressure and stirring speed are recorded by a Space View data acquisition system. The total volume of carbohydrate aqueous solution used was 10mL and the charged weight of catalyst was about 20mg. The catalytic reactions were carried out for the desired time or temperature without any base addition. The oxygen pressure was of 15bar in all experiments |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
86% | In methanol at 20℃; for 1.5h; | 4 Acetamido Cetyl Dimethyl Ammonium Glucuronate (CDA-Gluc) Example 4 Acetamido Cetyl Dimethyl Ammonium Glucuronate (CDA-Gluc) Glucuronic acid (100 mg, 0.515 mmol) and CDAOH (177 mg, 0.515 mmol) were combined in 25 mL of anhydrous methanol and stirred for 1.5 hours at room temperature. Methanol was removed by rotary evaporation and chloroform was added to dissolve the crude product. The solution was dried by stirring for a few minutes over anhydrous sodium sulfate following which the chloroform solution was decanted into a separate vessel, filtered and then evaporated to dryness. The initially gel-like product was triturated under ether to form a white powder that was isolated after pouring of the ether and drying the product under vacuum. Yield: 230.66 mg, 86%. |
Yield | Reaction Conditions | Operation in experiment |
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With glucuronic acid hydrolyase type HP-2 from Helixpomatia In aq. phosphate buffer at 37℃; for 5h; Enzymatic reaction; | 3.4. Enzymatic hydrolysis and absolute configuration determination General procedure: Compounds 1 (1.0 mg), 2 (4.4 mg), 3 (5.8 mg), 4 (3.2 mg), and 5(3.7 mg) were mixed with 0.2 mL KH2PO4/KOH buffer (pH=4.99) and0.1 mL glucuronic acid hydrolyase solution (Type HP-2 from Helixpomatia; Sigma-Aldrich), respectively. Then each solution was virtexmixed, and maintained for 5 h at 37 °C. After stopping, each reactionmixture was first extracted with CH2Cl2 to obtain the aglycone forfurther analysis, and then MeOH-CH3CN (2:1) was added to precipitatethe protein. After centrifugation for 5 min at 13,500 rpm, the uppersolution was analyzed using TLC over silica gel (EtOAc-EtOH-H2OHCOOH,6:4:1:1) by comparing with authentic sugars, proving thepresence of glucose (Glc) and glucuronic acid (GluA) in compounds1-5. Then the sugar residues was combined and subjected to CC onODS-A eluted with H2O to yield Glc and GluA. The optical rotationvalues of [α]D28+14.7 (c 0.19, H2O) for Glc and [α]D28+10.9 (c 0.35,H2O) for GluA confirmed the D-configurations of Glc and GluA.The CH2Cl2 extract was evaporated to yield the aglycones of 2-5,respectively, which were obtained as oils and smelled aromatic at roomtemperature. The 3R configurations for 2-4 and 2R configuration for 5were defined on the basis of the rotation value of [α]D30 + 6.7 (c 0.27,CH2Cl2) for the aglycone of 2, [α]D30-3.8 (c 0.16, CH2Cl2) for theaglycone of 3, [α]D30-4.0 (c 0.10, CH2Cl2) for the aglycone of 4, and [α]D30-5.5 (c 0.11, CH2Cl2) for the aglycone of 5. |
Yield | Reaction Conditions | Operation in experiment |
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With glucuronic acid hydrolyase type HP-2 from Helixpomatia In aq. phosphate buffer at 37℃; for 5h; Enzymatic reaction; | 3.4. Enzymatic hydrolysis and absolute configuration determination General procedure: Compounds 1 (1.0 mg), 2 (4.4 mg), 3 (5.8 mg), 4 (3.2 mg), and 5(3.7 mg) were mixed with 0.2 mL KH2PO4/KOH buffer (pH=4.99) and0.1 mL glucuronic acid hydrolyase solution (Type HP-2 from Helixpomatia; Sigma-Aldrich), respectively. Then each solution was virtexmixed, and maintained for 5 h at 37 °C. After stopping, each reactionmixture was first extracted with CH2Cl2 to obtain the aglycone forfurther analysis, and then MeOH-CH3CN (2:1) was added to precipitatethe protein. After centrifugation for 5 min at 13,500 rpm, the uppersolution was analyzed using TLC over silica gel (EtOAc-EtOH-H2OHCOOH,6:4:1:1) by comparing with authentic sugars, proving thepresence of glucose (Glc) and glucuronic acid (GluA) in compounds1-5. Then the sugar residues was combined and subjected to CC onODS-A eluted with H2O to yield Glc and GluA. The optical rotationvalues of [α]D28+14.7 (c 0.19, H2O) for Glc and [α]D28+10.9 (c 0.35,H2O) for GluA confirmed the D-configurations of Glc and GluA.The CH2Cl2 extract was evaporated to yield the aglycones of 2-5,respectively, which were obtained as oils and smelled aromatic at roomtemperature. The 3R configurations for 2-4 and 2R configuration for 5were defined on the basis of the rotation value of [α]D30 + 6.7 (c 0.27,CH2Cl2) for the aglycone of 2, [α]D30-3.8 (c 0.16, CH2Cl2) for theaglycone of 3, [α]D30-4.0 (c 0.10, CH2Cl2) for the aglycone of 4, and [α]D30-5.5 (c 0.11, CH2Cl2) for the aglycone of 5. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With glucuronic acid hydrolyase type HP-2 from Helixpomatia In aq. phosphate buffer at 37℃; for 5h; Enzymatic reaction; | 3.4. Enzymatic hydrolysis and absolute configuration determination General procedure: Compounds 1 (1.0 mg), 2 (4.4 mg), 3 (5.8 mg), 4 (3.2 mg), and 5(3.7 mg) were mixed with 0.2 mL KH2PO4/KOH buffer (pH=4.99) and0.1 mL glucuronic acid hydrolyase solution (Type HP-2 from Helixpomatia; Sigma-Aldrich), respectively. Then each solution was virtexmixed, and maintained for 5 h at 37 °C. After stopping, each reactionmixture was first extracted with CH2Cl2 to obtain the aglycone forfurther analysis, and then MeOH-CH3CN (2:1) was added to precipitatethe protein. After centrifugation for 5 min at 13,500 rpm, the uppersolution was analyzed using TLC over silica gel (EtOAc-EtOH-H2OHCOOH,6:4:1:1) by comparing with authentic sugars, proving thepresence of glucose (Glc) and glucuronic acid (GluA) in compounds1-5. Then the sugar residues was combined and subjected to CC onODS-A eluted with H2O to yield Glc and GluA. The optical rotationvalues of [α]D28+14.7 (c 0.19, H2O) for Glc and [α]D28+10.9 (c 0.35,H2O) for GluA confirmed the D-configurations of Glc and GluA.The CH2Cl2 extract was evaporated to yield the aglycones of 2-5,respectively, which were obtained as oils and smelled aromatic at roomtemperature. The 3R configurations for 2-4 and 2R configuration for 5were defined on the basis of the rotation value of [α]D30 + 6.7 (c 0.27,CH2Cl2) for the aglycone of 2, [α]D30-3.8 (c 0.16, CH2Cl2) for theaglycone of 3, [α]D30-4.0 (c 0.10, CH2Cl2) for the aglycone of 4, and [α]D30-5.5 (c 0.11, CH2Cl2) for the aglycone of 5. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With glucuronic acid hydrolyase type HP-2 from Helixpomatia In aq. phosphate buffer at 37℃; for 5h; Enzymatic reaction; | 3.4. Enzymatic hydrolysis and absolute configuration determination General procedure: Compounds 1 (1.0 mg), 2 (4.4 mg), 3 (5.8 mg), 4 (3.2 mg), and 5(3.7 mg) were mixed with 0.2 mL KH2PO4/KOH buffer (pH=4.99) and0.1 mL glucuronic acid hydrolyase solution (Type HP-2 from Helixpomatia; Sigma-Aldrich), respectively. Then each solution was virtexmixed, and maintained for 5 h at 37 °C. After stopping, each reactionmixture was first extracted with CH2Cl2 to obtain the aglycone forfurther analysis, and then MeOH-CH3CN (2:1) was added to precipitatethe protein. After centrifugation for 5 min at 13,500 rpm, the uppersolution was analyzed using TLC over silica gel (EtOAc-EtOH-H2OHCOOH,6:4:1:1) by comparing with authentic sugars, proving thepresence of glucose (Glc) and glucuronic acid (GluA) in compounds1-5. Then the sugar residues was combined and subjected to CC onODS-A eluted with H2O to yield Glc and GluA. The optical rotationvalues of [α]D28+14.7 (c 0.19, H2O) for Glc and [α]D28+10.9 (c 0.35,H2O) for GluA confirmed the D-configurations of Glc and GluA.The CH2Cl2 extract was evaporated to yield the aglycones of 2-5,respectively, which were obtained as oils and smelled aromatic at roomtemperature. The 3R configurations for 2-4 and 2R configuration for 5were defined on the basis of the rotation value of [α]D30 + 6.7 (c 0.27,CH2Cl2) for the aglycone of 2, [α]D30-3.8 (c 0.16, CH2Cl2) for theaglycone of 3, [α]D30-4.0 (c 0.10, CH2Cl2) for the aglycone of 4, and [α]D30-5.5 (c 0.11, CH2Cl2) for the aglycone of 5. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With glucuronic acid hydrolyase type HP-2 from Helixpomatia In aq. phosphate buffer at 37℃; for 5h; Enzymatic reaction; | 3.4. Enzymatic hydrolysis and absolute configuration determination General procedure: Compounds 1 (1.0 mg), 2 (4.4 mg), 3 (5.8 mg), 4 (3.2 mg), and 5(3.7 mg) were mixed with 0.2 mL KH2PO4/KOH buffer (pH=4.99) and0.1 mL glucuronic acid hydrolyase solution (Type HP-2 from Helixpomatia; Sigma-Aldrich), respectively. Then each solution was virtexmixed, and maintained for 5 h at 37 °C. After stopping, each reactionmixture was first extracted with CH2Cl2 to obtain the aglycone forfurther analysis, and then MeOH-CH3CN (2:1) was added to precipitatethe protein. After centrifugation for 5 min at 13,500 rpm, the uppersolution was analyzed using TLC over silica gel (EtOAc-EtOH-H2OHCOOH,6:4:1:1) by comparing with authentic sugars, proving thepresence of glucose (Glc) and glucuronic acid (GluA) in compounds1-5. Then the sugar residues was combined and subjected to CC onODS-A eluted with H2O to yield Glc and GluA. The optical rotationvalues of [α]D28+14.7 (c 0.19, H2O) for Glc and [α]D28+10.9 (c 0.35,H2O) for GluA confirmed the D-configurations of Glc and GluA.The CH2Cl2 extract was evaporated to yield the aglycones of 2-5,respectively, which were obtained as oils and smelled aromatic at roomtemperature. The 3R configurations for 2-4 and 2R configuration for 5were defined on the basis of the rotation value of [α]D30 + 6.7 (c 0.27,CH2Cl2) for the aglycone of 2, [α]D30-3.8 (c 0.16, CH2Cl2) for theaglycone of 3, [α]D30-4.0 (c 0.10, CH2Cl2) for the aglycone of 4, and [α]D30-5.5 (c 0.11, CH2Cl2) for the aglycone of 5. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride In 1,4-dioxane; water at 80℃; for 5h; | Acid hydrolysis and sugar identification Compound 1 (4 mg) was dissolved in 1.0 N HCl (Dioxane/H2O, 1:1) for 5 h in a water bath maintained at 80 0C. After cooling, the solution was evaporated under vacuum in a water bath at 60 0C and separated by solvent-solvent partition using CH2Cl2 and water. The sugar components in the aqueous layer were analyzed by silica gel TLC and compared with standard sugars. The solvent system was CH2Cl2/MeOH/H2O 2/1/0.2, and spots were visualized by spraying with 95% EtOH H2SO4 anisaldehyde (9/0.5/0.5, v/v), then heated at 200 0C for 7 min. The Rf values of glucuronic acid, galactose, and rhamnose on TLC were 0.15, 0.32, and 0.75, respectively. The sugar residue was dissolved in 3 mL of pyridine and heated with 10 mg of L-cysteine methyl ester at 80 0C for 60 min, before adding isothiocyanate (0.1 mL) to the reaction mixture and further incubated at 60 0C for 60 min as previously described (Tanaka et al. 2007). The reaction mixture was evaporated under vacuum. The reaction mixtures were directly analyzed by reversed-phase HPLC (Phenomenex column 250 ×4.6 mm, 5 μm), with a mobile phase MeCN-H2O (30-70); flow rate of 0.5 min/mL; UV detection at 250 nm. Peaks at tR (min) 10.2, 11.9, and 20.1 for D-glucuronic acid, D-galactose, and L-rhamnose, respectively. The retention times were compared to the standards with co-condition HPLC, and the absolute configurations of sugars from hydrolysis experiment were established. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride; water In 1,4-dioxane at 95℃; for 1h; Inert atmosphere; | Acid Hydrolysis of 1 and 2 Compounds 1 (2.5 mg) and2 (7.9 mg) were independently dissolved in 1 M HCl (dioxane-H2O, 1 : 1, 2.0 mL) and were heated at 95°C for 1 h underan Ar atmosphere. After dilution of the reaction mixture withH2O (5 mL), it was extracted with EtOAc (10 mL × 3). TheH2O residue was neutralized by the addition of Ag2CO3. Themixture was filtered and then passed through a Sep-Pak C18cartridge (Waters, Milford, MA, U.S.A.) eluted with H2O(5 mL × 3) to give sugar fractions (1.0 mg from 1, and 3.3 mgfrom 2). The sugar fractions were each analyzed by HPLC under the following conditions: 1) column, Aminex HPX-87H(7.8 mm i.d. × 300 mm, 5 μm, Bio-Rad Laboratories, CA,U.S.A.); solvent, 5 mM H2SO4 in H2O; flow rate, 0.6 mL/min;detection, tR (min) and optical rotation (OR). D-Glucuronicacid was detected in the sugar fractions obtained from 1 and2 (tR 8.23, positive optical rotation). 2) column, Capcell PakNH2 UG80 (4.6 mm i.d. × 250 mm, 5 μm, Shiseido, Tokyo,Japan); solvent, MeCN-H2O (17 : 3); flow rate, 1.0 mL/min;detection, tR (min) and OR. D-Galactose was detected in thesugar fractions obtained from 1 and 2 (tR 10.58, positive opticalrotation), and L-rhamnose in that from 2 (tR 6.58, negativeoptical rotation). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride; water In 1,4-dioxane at 95℃; for 1h; Inert atmosphere; | Acid Hydrolysis of 1 and 2 Compounds 1 (2.5 mg) and2 (7.9 mg) were independently dissolved in 1 M HCl (dioxane-H2O, 1 : 1, 2.0 mL) and were heated at 95°C for 1 h underan Ar atmosphere. After dilution of the reaction mixture withH2O (5 mL), it was extracted with EtOAc (10 mL × 3). TheH2O residue was neutralized by the addition of Ag2CO3. Themixture was filtered and then passed through a Sep-Pak C18cartridge (Waters, Milford, MA, U.S.A.) eluted with H2O(5 mL × 3) to give sugar fractions (1.0 mg from 1, and 3.3 mgfrom 2). The sugar fractions were each analyzed by HPLC under the following conditions: 1) column, Aminex HPX-87H(7.8 mm i.d. × 300 mm, 5 μm, Bio-Rad Laboratories, CA,U.S.A.); solvent, 5 mM H2SO4 in H2O; flow rate, 0.6 mL/min;detection, tR (min) and optical rotation (OR). D-Glucuronicacid was detected in the sugar fractions obtained from 1 and2 (tR 8.23, positive optical rotation). 2) column, Capcell PakNH2 UG80 (4.6 mm i.d. × 250 mm, 5 μm, Shiseido, Tokyo,Japan); solvent, MeCN-H2O (17 : 3); flow rate, 1.0 mL/min;detection, tR (min) and OR. D-Galactose was detected in thesugar fractions obtained from 1 and 2 (tR 10.58, positive opticalrotation), and L-rhamnose in that from 2 (tR 6.58, negativeoptical rotation). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
Stage #1: 3-O-[α-L-arabinofuranosyl(1→3)]-[β-D-galactopyranosyl(1→2)]-β-D-6-O-methylglucuro-nopyranosyl-21-O-angeloyl-22-O-(2-methyl)butyryl-R1-barrigenol With potassium hydroxide In 1,4-dioxane at 37℃; for 1h; Stage #2: With hydrogenchloride In water at 90℃; for 3h; | 4.4. Acid hydrolysis General procedure: Compounds (each 2.0 mg) were dissolved in 4M HCl (2 ml) and then heated in a H2O bath at 90 °C for 3 h to dryness, respectively. Mixture was extracted by CH2Cl2 and get CH2Cl2 extract. The water layer was analyzed by HPLC to detect tR (min): D-galactose: (+) 11.42 min, L-arabinose: (+) 9.25 min, D-glucose: (+) 10.67 min, L-rhamnoose:(-) 7.33 min. The HPLC analysis was carried using the colum of Kaseisorb LC NH2-60-5 (4.6 mmI.D.×250 mm) at room temperature with a flow rate of 0.8 ml/min. Mobile phase was 75%CH3CN/H2O. Optional detector: Shodex OR-2 (Range: 64; Atten: 2).The glucuronic acid was identified using GC method. Column: SupelucoSTBTM-1(30m×0.25 mmi.d.). Injetor temp.: 230. Detector temp.:230. Column temp.:230. He flow rate: 15 ml/min (methyl-D-glucuronicacid: 20.16 min). For compounds 1-2, 4-6, an alkaline hydrolysis experiment was performed prior to the aforementioned acid hydrolysis to remove either the methyl or the angeloyl group in the sugars (dissolvedin 1.5 ml 5% KOH: Dioxane 1.5 ml, stirred at 37 °C for 1 h). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride In water at 90℃; for 3h; | 4.4. Acid hydrolysis General procedure: Compounds (each 2.0 mg) were dissolved in 4M HCl (2 ml) and then heated in a H2O bath at 90 °C for 3 h to dryness, respectively. Mixture was extracted by CH2Cl2 and get CH2Cl2 extract. The water layer was analyzed by HPLC to detect tR (min): D-galactose: (+) 11.42 min, L-arabinose: (+) 9.25 min, D-glucose: (+) 10.67 min, L-rhamnoose:(-) 7.33 min. The HPLC analysis was carried using the colum of Kaseisorb LC NH2-60-5 (4.6 mmI.D.×250 mm) at room temperature with a flow rate of 0.8 ml/min. Mobile phase was 75%CH3CN/H2O. Optional detector: Shodex OR-2 (Range: 64; Atten: 2).The glucuronic acid was identified using GC method. Column: SupelucoSTBTM-1(30m×0.25 mmi.d.). Injetor temp.: 230. Detector temp.:230. Column temp.:230. He flow rate: 15 ml/min (methyl-D-glucuronicacid: 20.16 min). For compounds 1-2, 4-6, an alkaline hydrolysis experiment was performed prior to the aforementioned acid hydrolysis to remove either the methyl or the angeloyl group in the sugars (dissolvedin 1.5 ml 5% KOH: Dioxane 1.5 ml, stirred at 37 °C for 1 h). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
Stage #1: 3-O-β-D-6-O-methylglucuronopyranosyl-21-O-(3,4-di-O-angeloyl-β-D-fucopyranosyl) barrigenol C With potassium hydroxide In 1,4-dioxane at 37℃; for 1h; Stage #2: With hydrogenchloride; water at 90℃; for 3h; | 4.4. Acid hydrolysis General procedure: For compounds 4-7 (each 2.0 mg), an alkaline hydrolysis experiment (Brenner, 2013) was performed to remove the methyl group in sugars (dissolved in 1.5 ml 5% KOH:Dioxane 1.5 ml, stirred at 37 °C for 1 h). The powder afforded by the alkaline hydrolysis was dissolved in 4 M HCl (2 mL) and then heated in a H2O bath at 90 °C for 3 h to dryness. Mixture was extracted by CH2Cl2 and get CH2Cl2 extract. The water layer was identified using GC method by treating the residue with L-cysteine methyl ester hydrochloride (0.01 ml) in pyridine (0.02 ml) at 60 °C for 1 h. After this reaction, the solution was treated with N,O-bis (trimethyl silyl) trifluoroacetamide (0.01 ml) at 60 °C for 1 h. Column: Supelco STBTM-1 (30 m × 0.25 mmi.d.). Injetor temp.:230. Detector temp: 230 °C. Column temp.:230. He flow rate: 15 ml/min (methyl-Dglucuronic acid: 20.16 min). Additonally, residues were analyzed by HPLC with an NH2P-50 4 E column (4.6 × 250 mm, Showa Denko K.K., Japan) and an optical rotation detec-tor (JASCO ORD-4090, JASCO International Co, Ltd., Japan) to detect the configuration of the fucose using standard samples with the following condition: acetonitrile-water (75:25, v/v); flow rate, 0.8 mL/min. D-fucose, 9.5 min (positive). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
Stage #1: 3-O-β-D-6-O-methylglucuronopyranosyl-21-O-angeloyl-R1-barrigenol With potassium hydroxide In 1,4-dioxane at 37℃; for 1h; Stage #2: With hydrogenchloride; water at 90℃; for 3h; | 4.4. Acid hydrolysis General procedure: For compounds 4-7 (each 2.0 mg), an alkaline hydrolysis experiment (Brenner, 2013) was performed to remove the methyl group in sugars (dissolved in 1.5 ml 5% KOH:Dioxane 1.5 ml, stirred at 37 °C for 1 h). The powder afforded by the alkaline hydrolysis was dissolved in 4 M HCl (2 mL) and then heated in a H2O bath at 90 °C for 3 h to dryness. Mixture was extracted by CH2Cl2 and get CH2Cl2 extract. The water layer was identified using GC method by treating the residue with L-cysteine methyl ester hydrochloride (0.01 ml) in pyridine (0.02 ml) at 60 °C for 1 h. After this reaction, the solution was treated with N,O-bis (trimethyl silyl) trifluoroacetamide (0.01 ml) at 60 °C for 1 h. Column: Supelco STBTM-1 (30 m × 0.25 mmi.d.). Injetor temp.:230. Detector temp: 230 °C. Column temp.:230. He flow rate: 15 ml/min (methyl-Dglucuronic acid: 20.16 min). Additonally, residues were analyzed by HPLC with an NH2P-50 4 E column (4.6 × 250 mm, Showa Denko K.K., Japan) and an optical rotation detec-tor (JASCO ORD-4090, JASCO International Co, Ltd., Japan) to detect the configuration of the fucose using standard samples with the following condition: acetonitrile-water (75:25, v/v); flow rate, 0.8 mL/min. D-fucose, 9.5 min (positive). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
Stage #1: 3-O-β-D-6-O-methylglucuronopyranosyl-21-O-angeloyl-22-O-isobutyryl-R1-barrigenol With potassium hydroxide In 1,4-dioxane at 37℃; for 1h; Stage #2: With hydrogenchloride; water at 90℃; for 3h; | 4.4. Acid hydrolysis General procedure: For compounds 4-7 (each 2.0 mg), an alkaline hydrolysis experiment (Brenner, 2013) was performed to remove the methyl group in sugars (dissolved in 1.5 ml 5% KOH:Dioxane 1.5 ml, stirred at 37 °C for 1 h). The powder afforded by the alkaline hydrolysis was dissolved in 4 M HCl (2 mL) and then heated in a H2O bath at 90 °C for 3 h to dryness. Mixture was extracted by CH2Cl2 and get CH2Cl2 extract. The water layer was identified using GC method by treating the residue with L-cysteine methyl ester hydrochloride (0.01 ml) in pyridine (0.02 ml) at 60 °C for 1 h. After this reaction, the solution was treated with N,O-bis (trimethyl silyl) trifluoroacetamide (0.01 ml) at 60 °C for 1 h. Column: Supelco STBTM-1 (30 m × 0.25 mmi.d.). Injetor temp.:230. Detector temp: 230 °C. Column temp.:230. He flow rate: 15 ml/min (methyl-Dglucuronic acid: 20.16 min). Additonally, residues were analyzed by HPLC with an NH2P-50 4 E column (4.6 × 250 mm, Showa Denko K.K., Japan) and an optical rotation detec-tor (JASCO ORD-4090, JASCO International Co, Ltd., Japan) to detect the configuration of the fucose using standard samples with the following condition: acetonitrile-water (75:25, v/v); flow rate, 0.8 mL/min. D-fucose, 9.5 min (positive). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
Stage #1: 3-O-β-D-6-O-methylglucuronopyranosyl-21,22-di-O-angeloyl-R1-barrigenol With potassium hydroxide In 1,4-dioxane at 37℃; for 1h; Stage #2: With hydrogenchloride; water at 90℃; for 3h; | 4.4. Acid hydrolysis General procedure: For compounds 4-7 (each 2.0 mg), an alkaline hydrolysis experiment (Brenner, 2013) was performed to remove the methyl group in sugars (dissolved in 1.5 ml 5% KOH:Dioxane 1.5 ml, stirred at 37 °C for 1 h). The powder afforded by the alkaline hydrolysis was dissolved in 4 M HCl (2 mL) and then heated in a H2O bath at 90 °C for 3 h to dryness. Mixture was extracted by CH2Cl2 and get CH2Cl2 extract. The water layer was identified using GC method by treating the residue with L-cysteine methyl ester hydrochloride (0.01 ml) in pyridine (0.02 ml) at 60 °C for 1 h. After this reaction, the solution was treated with N,O-bis (trimethyl silyl) trifluoroacetamide (0.01 ml) at 60 °C for 1 h. Column: Supelco STBTM-1 (30 m × 0.25 mmi.d.). Injetor temp.:230. Detector temp: 230 °C. Column temp.:230. He flow rate: 15 ml/min (methyl-Dglucuronic acid: 20.16 min). Additonally, residues were analyzed by HPLC with an NH2P-50 4 E column (4.6 × 250 mm, Showa Denko K.K., Japan) and an optical rotation detec-tor (JASCO ORD-4090, JASCO International Co, Ltd., Japan) to detect the configuration of the fucose using standard samples with the following condition: acetonitrile-water (75:25, v/v); flow rate, 0.8 mL/min. D-fucose, 9.5 min (positive). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
Stage #1: 3-O-β-D-xylopyranosyl-(1→2)-β-D-glucuronopyranosylsophoradiol With acetic acid at 80℃; for 1h; Stage #2: With naphtho[2,3-d]imidazole; iodine at 20℃; for 1h; | Determination of monosaccharides of saponin derivatives General procedure: Compounds 1-5 (0.5mg each) were hydrolyzed by 0.25mL of H reagent (acetic acid) for 60minat 80°C. The mixture was then dried down. After that, added 1.5mg of A (naphthimidazole) and 0.5mL of B (iodine) reagent to the reaction vial, mixed well, stirred for 60minat room temperature. The reaction solution was dried down. Transferred the residue with 1.0mL of dd-water to an Eppendorf (1.5mL) and spun down the insoluble (3000rpm, 10min) (Lin et al., 2010), collected the supernatant. Similarly, the sugar standards were prepared without hydrolyzing. The supernatant was analyzed by using UPLC-MS. The flow rate of mobile phase A (ammonium formate 50mM in dd-water) and B (methanol) was 0.2mL/min. The injected volume was 5.0μL. The gradient program was set as follows: 0-10min, 35-50% B; 10-15min, isocratic elution with 50% B; 15-20min, 50-70% B. The column was washed with 100% of methanol for 5min before being re-equilibrated for 5min. The mass spectrometer was equipped with an electrospray interface controlled by the Xcalibur software (version 2.0, Thermo Fisher Scientific, Bremen, Germany) with two types of operations: The positive ion and negative ion modes. The ESI source was set at these parameters: Spray voltage of 3.5kV for the positive ion mode and -3.2kV for the negative ion mode; the capillary temperature was at 360°C and the source heater temperature was maintained at 350°C. During the analysis, the mass spectrometer performed high-solution (resolving power, r=15,000) full scan cycles (m/z 100-1500). The peaks of D-glucuronic acid - NAIM (Rt=5.28min) and D-xylose - NAIM (Rt=6.72min) were detected in the hydrolysate of all compounds 1-5. The peak of D-glucose - NAIM (Rt=6.22min) was only recorded in the hydrolysate of compound 1. The peaks of L-rhamnose - NAIM were detected in the hydrolysate of compounds 1-4at the retention time 7.50, 7.77, 8.00, and 8.59min, respectively. Whereas, the peaks of D-digitoxose - NAIM were exposed in the hydrolysate of compounds 1 and 3at 9.77 and 10,35min, respectively. These peaks were compared with the reference peaks. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride In water at 100℃; for 1h; | 2.4. Acid hydrolysis of triterpenoid saponins from P. glaucus General procedure: Compounds 1-13 (each 0.7 mg) were hydrolyzed with 0.1 mL of 0.5M HCl at 100 for 1 h. The solution was neutralized with Na2CO3solution and dried in vacuo. The residue was dissolved in 100 μL of Lcysteinemethyl ester hydrochloride in pyridine (5 mg/mL). Afterreacting for 1 h at 60 , 100 μL of trimethylsilylimidazole was added,and the mixture was left to react for 1 h at 60 . The supernatant wasanalyzed by HPLC based on a modified Tanaka’s method for determinationof sugar configuration [20]. The sugar derivatives showed retention times of 12.69, 12.80, 14.40, 15.09, and 22.28 min and wereidentified by comparison with those of the trimethylsilyl-L-cysteine derivativesof authentic D-glucose, D-glucuronic acid, L-arabinose, D-xylose,and L-rhamnose, respectively (Fig. S41, Supporting Information). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride In water at 100℃; for 1h; | 2.4. Acid hydrolysis of triterpenoid saponins from P. glaucus General procedure: Compounds 1-13 (each 0.7 mg) were hydrolyzed with 0.1 mL of 0.5M HCl at 100 for 1 h. The solution was neutralized with Na2CO3solution and dried in vacuo. The residue was dissolved in 100 μL of Lcysteinemethyl ester hydrochloride in pyridine (5 mg/mL). Afterreacting for 1 h at 60 , 100 μL of trimethylsilylimidazole was added,and the mixture was left to react for 1 h at 60 . The supernatant wasanalyzed by HPLC based on a modified Tanaka’s method for determinationof sugar configuration [20]. The sugar derivatives showed retention times of 12.69, 12.80, 14.40, 15.09, and 22.28 min and wereidentified by comparison with those of the trimethylsilyl-L-cysteine derivativesof authentic D-glucose, D-glucuronic acid, L-arabinose, D-xylose,and L-rhamnose, respectively (Fig. S41, Supporting Information). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride In water at 100℃; for 1h; | 2.4. Acid hydrolysis of triterpenoid saponins from P. glaucus General procedure: Compounds 1-13 (each 0.7 mg) were hydrolyzed with 0.1 mL of 0.5M HCl at 100 for 1 h. The solution was neutralized with Na2CO3solution and dried in vacuo. The residue was dissolved in 100 μL of Lcysteinemethyl ester hydrochloride in pyridine (5 mg/mL). Afterreacting for 1 h at 60 , 100 μL of trimethylsilylimidazole was added,and the mixture was left to react for 1 h at 60 . The supernatant wasanalyzed by HPLC based on a modified Tanaka’s method for determinationof sugar configuration [20]. The sugar derivatives showed retention times of 12.69, 12.80, 14.40, 15.09, and 22.28 min and wereidentified by comparison with those of the trimethylsilyl-L-cysteine derivativesof authentic D-glucose, D-glucuronic acid, L-arabinose, D-xylose,and L-rhamnose, respectively (Fig. S41, Supporting Information). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride In water at 100℃; for 1h; | 2.4. Acid hydrolysis of triterpenoid saponins from P. glaucus General procedure: Compounds 1-13 (each 0.7 mg) were hydrolyzed with 0.1 mL of 0.5M HCl at 100 for 1 h. The solution was neutralized with Na2CO3solution and dried in vacuo. The residue was dissolved in 100 μL of Lcysteinemethyl ester hydrochloride in pyridine (5 mg/mL). Afterreacting for 1 h at 60 , 100 μL of trimethylsilylimidazole was added,and the mixture was left to react for 1 h at 60 . The supernatant wasanalyzed by HPLC based on a modified Tanaka’s method for determinationof sugar configuration [20]. The sugar derivatives showed retention times of 12.69, 12.80, 14.40, 15.09, and 22.28 min and wereidentified by comparison with those of the trimethylsilyl-L-cysteine derivativesof authentic D-glucose, D-glucuronic acid, L-arabinose, D-xylose,and L-rhamnose, respectively (Fig. S41, Supporting Information). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride In water at 100℃; for 1h; | 2.4. Acid hydrolysis of triterpenoid saponins from P. glaucus General procedure: Compounds 1-13 (each 0.7 mg) were hydrolyzed with 0.1 mL of 0.5M HCl at 100 for 1 h. The solution was neutralized with Na2CO3solution and dried in vacuo. The residue was dissolved in 100 μL of Lcysteinemethyl ester hydrochloride in pyridine (5 mg/mL). Afterreacting for 1 h at 60 , 100 μL of trimethylsilylimidazole was added,and the mixture was left to react for 1 h at 60 . The supernatant wasanalyzed by HPLC based on a modified Tanaka’s method for determinationof sugar configuration [20]. The sugar derivatives showed retention times of 12.69, 12.80, 14.40, 15.09, and 22.28 min and wereidentified by comparison with those of the trimethylsilyl-L-cysteine derivativesof authentic D-glucose, D-glucuronic acid, L-arabinose, D-xylose,and L-rhamnose, respectively (Fig. S41, Supporting Information). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With hydrogenchloride In water at 100℃; for 1h; | 2.4. Acid hydrolysis of triterpenoid saponins from P. glaucus General procedure: Compounds 1-13 (each 0.7 mg) were hydrolyzed with 0.1 mL of 0.5M HCl at 100 for 1 h. The solution was neutralized with Na2CO3solution and dried in vacuo. The residue was dissolved in 100 μL of Lcysteinemethyl ester hydrochloride in pyridine (5 mg/mL). Afterreacting for 1 h at 60 , 100 μL of trimethylsilylimidazole was added,and the mixture was left to react for 1 h at 60 . The supernatant wasanalyzed by HPLC based on a modified Tanaka’s method for determinationof sugar configuration [20]. The sugar derivatives showed retention times of 12.69, 12.80, 14.40, 15.09, and 22.28 min and wereidentified by comparison with those of the trimethylsilyl-L-cysteine derivativesof authentic D-glucose, D-glucuronic acid, L-arabinose, D-xylose,and L-rhamnose, respectively (Fig. S41, Supporting Information). |
Tags: 6556-12-3 synthesis path| 6556-12-3 SDS| 6556-12-3 COA| 6556-12-3 purity| 6556-12-3 application| 6556-12-3 NMR| 6556-12-3 COA| 6556-12-3 structure
Precautionary Statements-General | |
Code | Phrase |
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Prevention | |
Code | Phrase |
P201 | Obtain special instructions before use. |
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P264 | Wash hands thoroughly after handling. |
P265 | Wash skin thouroughly after handling. |
P270 | Do not eat, drink or smoke when using this product. |
P271 | Use only outdoors or in a well-ventilated area. |
P272 | Contaminated work clothing should not be allowed out of the workplace. |
P273 | Avoid release to the environment. |
P280 | Wear protective gloves/protective clothing/eye protection/face protection. |
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P282 | Wear cold insulating gloves/face shield/eye protection. |
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Response | |
Code | Phrase |
P301 | IF SWALLOWED: |
P304 | IF INHALED: |
P305 | IF IN EYES: |
P306 | IF ON CLOTHING: |
P307 | IF exposed: |
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P320 | |
P302 + P352 | IF ON SKIN: wash with plenty of soap and water. |
P321 | |
P322 | |
P330 | Rinse mouth. |
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P332 | IF SKIN irritation occurs: |
P333 | If skin irritation or rash occurs: |
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P335 | Brush off loose particles from skin. |
P336 | Thaw frosted parts with lukewarm water. Do not rub affected area. |
P337 | If eye irritation persists: |
P338 | Remove contact lenses, if present and easy to do. Continue rinsing. |
P340 | Remove victim to fresh air and keep at rest in a position comfortable for breathing. |
P341 | If breathing is difficult, remove victim to fresh air and keep at rest in a position comfortable for breathing. |
P342 | If experiencing respiratory symptoms: |
P350 | Gently wash with plenty of soap and water. |
P351 | Rinse cautiously with water for several minutes. |
P352 | Wash with plenty of soap and water. |
P353 | Rinse skin with water/shower. |
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P362 | Take off contaminated clothing and wash before reuse. |
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P370 | In case of fire: |
P371 | In case of major fire and large quantities: |
P372 | Explosion risk in case of fire. |
P373 | DO NOT fight fire when fire reaches explosives. |
P374 | Fight fire with normal precautions from a reasonable distance. |
P376 | Stop leak if safe to do so. Oxidising gases (section 2.4) 1 |
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P378 | |
P380 | Evacuate area. |
P381 | Eliminate all ignition sources if safe to do so. |
P390 | Absorb spillage to prevent material damage. |
P391 | Collect spillage. Hazardous to the aquatic environment |
P301 + P310 | IF SWALLOWED: Immediately call a POISON CENTER or doctor/physician. |
P301 + P312 | IF SWALLOWED: call a POISON CENTER or doctor/physician IF you feel unwell. |
P301 + P330 + P331 | IF SWALLOWED: Rinse mouth. Do NOT induce vomiting. |
P302 + P334 | IF ON SKIN: Immerse in cool water/wrap in wet bandages. |
P302 + P350 | IF ON SKIN: Gently wash with plenty of soap and water. |
P303 + P361 + P353 | IF ON SKIN (or hair): Remove/Take off Immediately all contaminated clothing. Rinse SKIN with water/shower. |
P304 + P312 | IF INHALED: Call a POISON CENTER or doctor/physician if you feel unwell. |
P304 + P340 | IF INHALED: Remove victim to fresh air and Keep at rest in a position comfortable for breathing. |
P304 + P341 | IF INHALED: If breathing is difficult, remove victim to fresh air and keep at rest in a position comfortable for breathing. |
P305 + P351 + P338 | IF IN EYES: Rinse cautiously with water for several minutes. Remove contact lenses, if present and easy to do. Continue rinsing. |
P306 + P360 | IF ON CLOTHING: Rinse Immediately contaminated CLOTHING and SKIN with plenty of water before removing clothes. |
P307 + P311 | IF exposed: call a POISON CENTER or doctor/physician. |
P308 + P313 | IF exposed or concerned: Get medical advice/attention. |
P309 + P311 | IF exposed or if you feel unwell: call a POISON CENTER or doctor/physician. |
P332 + P313 | IF SKIN irritation occurs: Get medical advice/attention. |
P333 + P313 | IF SKIN irritation or rash occurs: Get medical advice/attention. |
P335 + P334 | Brush off loose particles from skin. Immerse in cool water/wrap in wet bandages. |
P337 + P313 | IF eye irritation persists: Get medical advice/attention. |
P342 + P311 | IF experiencing respiratory symptoms: call a POISON CENTER or doctor/physician. |
P370 + P376 | In case of fire: Stop leak if safe to Do so. |
P370 + P378 | In case of fire: |
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P370 + P380 + P375 | In case of fire: Evacuate area. Fight fire remotely due to the risk of explosion. |
P371 + P380 + P375 | In case of major fire and large quantities: Evacuate area. Fight fire remotely due to the risk of explosion. |
Storage | |
Code | Phrase |
P401 | |
P402 | Store in a dry place. |
P403 | Store in a well-ventilated place. |
P404 | Store in a closed container. |
P405 | Store locked up. |
P406 | Store in corrosive resistant/ container with a resistant inner liner. |
P407 | Maintain air gap between stacks/pallets. |
P410 | Protect from sunlight. |
P411 | |
P412 | Do not expose to temperatures exceeding 50 oC/ 122 oF. |
P413 | |
P420 | Store away from other materials. |
P422 | |
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P410 + P412 | Protect from sunlight. Do not expose to temperatures exceeding 50 oC/122oF. |
P411 + P235 | Keep cool. |
Disposal | |
Code | Phrase |
P501 | Dispose of contents/container to ... |
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Physical hazards | |
Code | Phrase |
H200 | Unstable explosive |
H201 | Explosive; mass explosion hazard |
H202 | Explosive; severe projection hazard |
H203 | Explosive; fire, blast or projection hazard |
H204 | Fire or projection hazard |
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H221 | Flammable gas |
H222 | Extremely flammable aerosol |
H223 | Flammable aerosol |
H224 | Extremely flammable liquid and vapour |
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H228 | Flammable solid |
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H230 | May react explosively even in the absence of air |
H231 | May react explosively even in the absence of air at elevated pressure and/or temperature |
H240 | Heating may cause an explosion |
H241 | Heating may cause a fire or explosion |
H242 | Heating may cause a fire |
H250 | Catches fire spontaneously if exposed to air |
H251 | Self-heating; may catch fire |
H252 | Self-heating in large quantities; may catch fire |
H260 | In contact with water releases flammable gases which may ignite spontaneously |
H261 | In contact with water releases flammable gas |
H270 | May cause or intensify fire; oxidizer |
H271 | May cause fire or explosion; strong oxidizer |
H272 | May intensify fire; oxidizer |
H280 | Contains gas under pressure; may explode if heated |
H281 | Contains refrigerated gas; may cause cryogenic burns or injury |
H290 | May be corrosive to metals |
Health hazards | |
Code | Phrase |
H300 | Fatal if swallowed |
H301 | Toxic if swallowed |
H302 | Harmful if swallowed |
H303 | May be harmful if swallowed |
H304 | May be fatal if swallowed and enters airways |
H305 | May be harmful if swallowed and enters airways |
H310 | Fatal in contact with skin |
H311 | Toxic in contact with skin |
H312 | Harmful in contact with skin |
H313 | May be harmful in contact with skin |
H314 | Causes severe skin burns and eye damage |
H315 | Causes skin irritation |
H316 | Causes mild skin irritation |
H317 | May cause an allergic skin reaction |
H318 | Causes serious eye damage |
H319 | Causes serious eye irritation |
H320 | Causes eye irritation |
H330 | Fatal if inhaled |
H331 | Toxic if inhaled |
H332 | Harmful if inhaled |
H333 | May be harmful if inhaled |
H334 | May cause allergy or asthma symptoms or breathing difficulties if inhaled |
H335 | May cause respiratory irritation |
H336 | May cause drowsiness or dizziness |
H340 | May cause genetic defects |
H341 | Suspected of causing genetic defects |
H350 | May cause cancer |
H351 | Suspected of causing cancer |
H360 | May damage fertility or the unborn child |
H361 | Suspected of damaging fertility or the unborn child |
H361d | Suspected of damaging the unborn child |
H362 | May cause harm to breast-fed children |
H370 | Causes damage to organs |
H371 | May cause damage to organs |
H372 | Causes damage to organs through prolonged or repeated exposure |
H373 | May cause damage to organs through prolonged or repeated exposure |
Environmental hazards | |
Code | Phrase |
H400 | Very toxic to aquatic life |
H401 | Toxic to aquatic life |
H402 | Harmful to aquatic life |
H410 | Very toxic to aquatic life with long-lasting effects |
H411 | Toxic to aquatic life with long-lasting effects |
H412 | Harmful to aquatic life with long-lasting effects |
H413 | May cause long-lasting harmful effects to aquatic life |
H420 | Harms public health and the environment by destroying ozone in the upper atmosphere |
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