Glycosylation of hesperetin by plant cell cultures

The biotransformation of hesperetin by cultured cells of Ipomoea batatas and Eucalyptus perriniana was investigated. Three glycosides, hesperetin 3'-O-beta-D-glucopyranoside (33 microg/g fr. wt of cells), hesperetin 3',7-O-beta-D-diglucopyranoside (217 microg/g fr. wt of cells), and hesperetin 7-O-[6-O-(beta-D-glucopyranosyl)]-beta-d-glucopyranoside (beta-gentiobioside, 22 microg/g fr. wt of cells), together with three hitherto known glycosides, hesperetin 5-O-beta-d-glucopyranoside (23 microg/g fr. wt of cells), hesperetin 7-O-beta-D-glucopyranoside (57 microg/g fr. wt of cells), and hesperetin 7-O-[6-O-(alpha-L-rhamnopyranosyl)]-beta-D-glucopyranoside (beta-rutinoside, hesperidin, 13 microg/g fr. wt of cells), were isolated from cultured suspension cells of E. perriniana that had been treated with hesperetin. Oligosaccharide chains were regioselectively formed at the C-7 position of hesperetin to afford beta-gentiobioside and beta-rutinoside. On the other hand, cultured I. batatas cells converted hesperetin into hesperetin 3'-O-beta-D-glucopyranoside (60 microg/g fr. wt of cells), hesperetin 5-O-beta-D-glucopyranoside (23 microg/g fr. wt of cells), and hesperetin 7-O-beta-D-glucopyranoside (110 microg/g fr. wt of cells).     

Biotransformation of thymol, carvacrol, and eugenol by cultured cells of Eucalyptus perriniana

The biotransformations of aroma compounds of spices, such as thymol (1), carvacrol (2), and eugenol (3), were investigated using cultured plant cells of Eucalyptus perriniana. Besides a beta-glucoside product (4, 3%), a biotransformation product, i.e., 5-methyl-2-(1-methylethyl)phenyl 6-O-(beta-d-glucopyranosyl)-beta-d-glucopyranoside (5, beta-gentiobioside, 87%), was isolated from the suspension cells after the five-day incubation of 1. On administration of 2, a beta-glucoside (6, 5%) and a beta-gentiobioside, i.e., 2-methyl-5-(1-methylethyl)phenyl 6-O-(beta-d-glucopyranosyl)-beta-d-glucopyranoside (7, 56%), were produced. Furthermore, E. perriniana cells also converted 3 into the corresponding beta-glucoside (8, 7%) and beta-gentiobioside (9, 58%). The cultured cells of E. perriniana are able to convert these aroma compounds of spices into glycosides which are accumulated in the cells.     

Glycosylation of tocopherols by cultured cells of Eucalyptus perriniana

Cultured suspension cells of Eucalyptus perriniana converted exogenously administered alpha-tocopherol into alpha-tocopheryl 6-O-beta-d-glucopyranoside (46mug/gfr. wt of cells) and two biotransformation products: alpha-tocopheryl 6-O-(6-O-beta-d-glucopyranosyl)-beta-d-glucopyranoside (19mug/gfr. wt of cells) and alpha-tocopheryl 6-O-(6-O-alpha-l-rhamnopyranosyl)-beta-d-glucopyranoside (6mug/gfr. wt of cells). On the other hand, two other compounds, i.e., delta-tocopheryl 6-O-(6-O-beta-d-glucopyranosyl)-beta-d-glucopyranoside (27mug/g fr. wt of cells) and delta-tocopheryl 6-O-(6-O-alpha-l-rhamnopyranosyl)-beta-d-glucopyranoside (12mug/g fr. wt of cells), together with delta-tocopheryl 6-O-beta-d-glucopyranoside (63mug/g fr. wt of cells) were isolated from suspension cells following the administration of delta-tocopherol.     

Acetylated flavonol diglucosides from Meconopsis quintuplinervia

Four acetylated flavonol diglucosides, quercetin 3-O-[2'-O-acetyl-beta-d-glucopyranosyl-(1-->6)-beta-d-glucopyranoside], quercetin 3-O-[2',6'-O-diacetyl-beta-d-glucopyranosyl-(1-->6)-beta-d-glucopyranoside], isorhamnetin 3-O-[2'-O-acetyl-beta-d-glucopyranosyl-(1-->6)-beta-d-glucopyranoside], and quercetin 3-O-[2'-O-acetyl-alpha-l-arabinopyranosyl-(1-->6)-beta-d-glucopyranoside], together with five known flavonol glycosides quercetin 3-O-beta-d-glucopyranoside, kaempferol 3-O-beta-d-glucopyranoside, quercetin 3-O-[beta-d-galactopyranosyl-(1-->6)-glucopyranoside], isorhamnetin 3-O-[beta-d-galactopyranosyl-(1-->6)-beta-d-glucopyranoside], and kaempferol 3-O-[beta-d-glucopyranosyl-(1-->2)-beta-d-glucopyranoside] have been isolated from Meconopsis quintuplinervia. Their structures were determined using chemical and spectroscopic methods including HRFABMS, (1)H-(1)H COSY, HSQC and HMBC experiments.     

Glycosylation of capsaicin and 8-nordihydrocapsaicin by cultured cells of Catharanthus roseus

The glycosylation of capsaicin and 8-nordihydrocapsaicin was investigated using cultured cells of Catharanthus roseus. In addition to capsaicin 4-O-beta-d-glucopyranoside (170 microg/g fr. wt of cells), the biotransformation products, capsaicin 4-O-(6-O-beta-D-xylopyranosyl)-beta-D-glucopyranoside (116 microg/g fr. wt of cells) and capsaicin 4-O-(6-O-alpha-L-arabinopyranosyl)-beta-D-glucopyranoside (83 microg/g fr. wt of cells), were isolated from the cell suspension after three days of incubation with capsaicin. Two other compounds, 8-nordihydrocapsaicin 4-O-(6-O-beta-D-xylopyranosyl)-beta-D-glucopyranoside (171 microg/g fr. wt of cells) and 8-nordihydrocapsaicin 4-O-(6-O-alpha-L-arabinopyranosyl)-beta-D-glucopyranoside (122 microg/g fr. wt of cells), together with the known 8-nordihydrocapsaicin 4-O-beta-D-glucopyranoside (204 microg/g fr. wt of cells) were also isolated from the cell suspension after incubation with 8-nordihydrocapsaicin.     

Steroidal glycosides from the rhizomes of Ruscus hypophyllum

Seven steroidal glycosides, along with one known glycoside, were isolated from the rhizomes of Ruscus hypophyllum (Liliaceae). Comprehensive spectroscopic analysis, including 2D NMR spectroscopy, and the results of acid hydrolysis allowed the chemical structures of the compounds to be assigned as (23S,25R)-23-hydroxyspirost-5-en-3beta-yl O-alpha-l-rhamnopyranosyl-(1-->4)-beta-d-glucopyranoside (1), 1beta-hydroxyspirosta-5,25(27)-dien-3beta-yl O-alpha-l-rhamnopyranosyl-(1-->4)-beta-d-glucopyranoside (2), (22S)-16beta,22-dihydroxycholest-5-en-3beta-yl O-alpha-l-rhamnopyranosyl-(1-->4)-beta-d-glucopyranoside (3), (22S)-16beta-[(beta-d-glucopyranosyl)oxy]-22-hydroxycholest-5-en-3beta-yl O-alpha-l-rhamnopyranosyl-(1-->4)-beta-d-glucopyranoside (4), (22S)-16beta-[(beta-d-glucopyranosyl)oxy]-22-hydroxycholest-5-en-3beta-yl beta-d-glucopyranoside (5), (22S)-16beta-[(beta-d-glucopyranosyl)oxy]-3beta,22-dihydroxycholest-5-en-1beta-yl O-alpha-l-rhamnopyranosyl-(1-->2)-(3,4-di-O-acetyl-beta-d-xylopyranoside) (6), and (22S)-16beta-[(beta-d-glucopyranosyl)oxy]-3beta,22-dihydroxycholest-5-en-1beta-yl O-alpha-l-rhamnopyranosyl-(1-->2)-O-[beta-d-xylopyranosyl-(1-->3)]-beta-d-xylopyranoside (7), respectively. This is the first isolation of a series of cholestane glycosides from a Ruscus species.     

Acylated cyanidin 3-sambubioside-5-glucosides in three garden plants of the Cruciferae

Seven acylated cyanidin 3-sambubioside-5-glucosides were isolated from the flowers of three garden plants in the Cruciferae. Specifically, four pigments were isolated from Lobularia maritima (L.) Desv., together with a known pigment, as well as, three pigments from Lunaria annua L., and two known pigments from Cheiranthus cheiri L. These pigments were determined to be cyanidin 3-O-[2-O-((acyl-II)-(beta-d-xylopyranosyl))-6-O-(acyl-I)-beta-d-glucopyranoside]-5-O-[6-O-(acyl-III)-beta-d-glucopyranoside], in which the acyl-I group is represented by glucosyl-p-coumaric acid, p-coumaric acid and ferulic acid, acyl-II by caffeic acid and ferulic acid, and acyl-III by malonic acid, respectively. The distribution and biosynthesis of acylated cyanidin 3-sambubioside-5-glucosides are discussed according to the variations of acylation and glucosylation at their 3-sambubiose residues.     

Steroidal saponins from roots of Asparagus officinalis

Sarsasapogenin M (1) and sarsasapogenin N (2), two new oligospirostanosides with a unique aglycone moiety, (25S)-5beta-spirostan-3beta, 17alpha-diol, along with seven known compounds (25S)-5beta-spirostan-3beta-ol-3-O-beta-d-glucopyranosyl-(1,2)-[beta-d-xylopyranosyl-(1,4)]-beta-d-glucopyranoside (3), (25S)-5beta-spirostan-3beta-ol-3-O-beta-d-glucopyranosyl-(1,2)-beta-d-glucopyranoside (4), (25S)-5beta-spirostan-3beta-ol-3-O-alpha-l-rhamnopyranosyl-(1,2)-[alpha-l-rhamnopyranosyl-(1,4)]-beta-d-glucopyranoside (5), (25S)26-O-beta-d-glucopyranosyl-5beta-furost-20 (22)-ene-3beta,26-diol-3-O-beta-d-glucopyranosyl-(1,2)-beta-d-glucopyranoside (6), yamogenin (7), beta-sitosterol (8), and sitosterol-beta-d-glucoside (9) were isolated from the roots of Asparagus officinalis L. Their structures were determined by spectral analysis, including extensive 1D and 2D NMR experiments.     

Glycosylation of sesamol by cultured plant cells

The glycosylation of sesamol was investigated using cultured cells of Nicotiana tabacum and Eucalyptus perriniana. The cultured suspension cells of N. tabacum converted sesamol into its β-glucoside (7%) as well as the disaccharide, sesamyl 6-O-(β-D-glucopyranosyl)-β-D-glucopyranoside (β-gentiobioside, 30%). On the other hand, sesamyl 6-O-(α-L-rhamnopyranosyl)-β-D-glucopyranoside (β-rutinoside, 56%), together with the β-glucoside (3%), was produced when sesamol was incubated with suspension cells of E. perriniana.     

Triterpenoid saponins from the fruits and galls of Sapindus mukorossi

Six saponins, sapinmusaponin K (1) [hederagenin-3-O-(3-O-acetyl-alpha-L-arabinopyranosyl)-(1-->3)-alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranoside], sapinmusaponin L (2) [hederagenin-3-O-(4-O-acetyl-alpha-L-arabinopyranosyl)-(1-->3)-alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabino-pyranoside], sapinmusaponin M (3) [hederagenin-3-O-(2,3-O-diacetyl-beta-D-xylopyranosyl)-(1-->3)-alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranoside], sapinmusaponin N (4) [hederagenin-3-O-(2,4-O-diacetyl-beta-D-xylopyranosyl)-(1-->3)-alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranoside], sapinmusaponin O (5) [3,7,20(S)-trihydroxydammar-24-ene-3-O-alpha-L-rhamnopyranosyl-(1-->2)-beta-D-glucopyranoside], and sapinmusaponin P (6) [3,7,20(R)-trihydroxydammar-24-ene-3-O-alpha-L-rhamnopyranosyl-(1-->2)-beta-d-glucopyranoside], along with seven known saponins (7-13), were isolated from fruits and the galls of Sapindus mukorossi. Their structures were elucidated by 1D and 2D NMR spectroscopic techniques and acid hydrolysis. Biological evaluation indicated that saponins 1-4 and 7-13 showed moderate cytotoxicity against several human tumor cell lines.     

In vitro enzymatic modification of puerarin to puerarin glycosides by maltogenic amylase

Puerarin (daidzein 8-C-glucoside), the most abundant isoflavone in Puerariae radix, is prescribed to treat coronary heart disease, cardiac infarction, problems in ocular blood flow, sudden deafness, and alcoholism. However, puerarin cannot be given by injection due to its low solubility in water. To increase its solubility, puerarin was transglycosylated using various enzymes. Bacillus stearothermophilus maltogenic amylase (BSMA) was the most effective transferase used compared with Thermotoga maritima maltosyl transferase (TMMT), Thermus scotoductus 4-alpha-glucanotransferase (TS4alphaGTase), and Bacillus sp. I-5 cyclodextrin glucanotransferase (BSCGTase). TMMT and TS4alphaGTase lacked acceptor specificity for puerarin, which lacks an O-glucoside linkage between D-glucose and 7-OH-daidzein. The yield exceeded 70% when reacting 1% puerarin (acceptor), 3.0% soluble starch (donor), and 5U/100 microL BSMA at 55 degrees C for 45 min. The two major transfer products of the BSMA reaction were purified using C(18) and GPC chromatography. Their structures were identified as alpha-d-glucosyl-(1-->6)-puerarin and alpha-D-maltosyl-(1-->6)-puerarin using ESI+ TOF MS-MS and 13C NMR spectroscopy. The solubility of the transfer products was 14 and 168 times higher than that of puerarin, respectively.     

Triacylated cyanidin 3-(3X-glucosylsambubioside)-5-glucosides from the flowers of Malcolmia maritima

Three acylated cyanidin 3-(3(X)-glucosylsambubioside)-5-glucosides (1-3) and one non-acylated cyanidin 3-(3(X)-glucosylsambubioside)-5-glucoside (4) were isolated from the purple-violet or violet flowers and purple stems of Malcolmia maritima (L.) R. Br (the Cruciferae), and their structures were determined by chemical and spectroscopic methods. In the flowers of this plant, pigment 1 was determined to be cyanidin 3-O-[2-O-(2-O-(trans-sinapoyl)-3-O-(beta-D-glucopyranosyl)-beta-D-xylopyranosyl)-6-O-(trans-p-coumaroyl)-beta-D-glucopyranoside]-5-O-[6-O-(malonyl)-(beta-D-glucopyranoside) as a major pigment, and a minor pigment 2 was determined to be the cis-p-coumaroyl isomer of pigment 1. In the stems, pigment 3 was determined to be cyanidin 3-O-[2-O-(2-O-(trans-sinapoyl)-3-O-(beta-D-glucopyranosyl)-beta-D-xylopyranosyl)-6-O-(trans-p-coumaroyl)-beta-d-glucopyranoside]-5-O-(beta-D-glucopyranoside) as a major anthocyanin, and also a non-acylated anthocyanin, cyanidin 3-O-[2-O-(3-O-(beta-D-glucopyranosyl)-beta-D-xylopyranosyl)-beta-D-glucopyranoside]-5-O-(beta-D-glucopyranoside) was determined to be a minor pigment (pigment 4). In this study, it was established that the acylation-enzymes of malonic acid has important roles for the acylation of 5-glucose residues of these anthocyanins in the flower-tissues of M. maritima; however, the similar enzymatic reactions seemed to be inhibited or lacking in the stem-tissues.     

Glucosylation of phenols and phenylalkyl alcohols using cultured plant cells

Cultured plant cells of Eucalyptus perriniana can convert phenol and phenylalkyl alcohols [C(6)H(5)(CH(2))(n)OH, n=0-3] into the corresponding beta-D-glucopyranosides in a good yield. The cells preferentially glucosylated phenylmethanol (n=1, 59% yield) rather than phenol (n=0, 49%), 2-phenylethanol (n=2, 38%), and 3-phenylpropan-1-ol (n=3, 20%). On the other hand, 2-, 3-, and 4-hydroxyphenylmethanols were also glucosylated to (hydroxymethyl)phenyl beta-D-glucopyranosides and (hydroxyphenyl)methyl beta-D-glucopyranosides by cultured E. perriniana cells.     

Saponins and acylated saponins from Dizygotheca kerchoveana

Four new triterpenoid saponins were isolated from the leaves and stem of branches of Dizygotheca kerchoveana along with seven known ones. The new saponins were respectively characterized as 3-O-[beta-D-glucopyranosyl-(1-->3)]-[beta-D-glucopyranosyl-(1-->2)]-alpha-L-arabinopyranosyl echinocystic acid, 3-O-[beta-D-glucopyranosyl-(1-->3)]-[beta-D-glucopyranosyl-(1-->2)]-alpha-L-arabinopyranosyl echinocystic acid 28-O-[alpha-L-rhamnopyranosyl-(1-->4)-beta-D-glucopyranosyl-(1-->6)-beta-D-glucopyranosyl] ester, 3-O-[beta-D-3-O-trans-p-coumaroyl-glucopyranosyl-(1-->3)]-[beta-D-glucopyranosyl-(1-->2)]-alpha-L-arabinopyranosyl echinocystic acid 28-O-[alpha-L-rhamnopyranosyl-(1-->4)-beta-D-glucopyranosyl-(1-->6)-beta-D-glucopyranosyl] ester and 3-O-[beta-d-3-O-cis-p-coumaroyl-glucopyranosyl-(1-->3)]-[beta-D-glucopyranosyl-(1-->2)]-alpha-L-arabinopyranosyl echinocystic acid 28-O-[alpha-L-rhamnopyranosyl-(1-->4)-beta-D-glucopyranosyl-(1-->6)-beta-D-glucopyranosyl] ester. Their structures were elucidated by 1D and 2D NMR experiments, FAB-MS as well as chemical means.     

Glycosides from Phlomis lunariifolia

An aliphatic alcohol glycoside, lunaroside 1-octen-3-yl [O-beta-apiofuranosyl-(1-->6)-O-[beta-glucopyranosyl-(1-->2)]-beta-glucopyranoside, a phenylethanoid glycoside, lunariifolioside 2-(3,4-dihydroxyphenyl)ethylO-beta-apiofuranosyl-(1-->6)-O-[O-beta-apiofuranosyl-(1-->4)-alpha-rhamnopyranosyl-(1-->3)]-4-O-(E)-caffeoyl-beta-glucopyranoside and a flavone glycoside, luteolin 7-O-[4-O-acetyl-alpha-rhamnopyranosyl-(1-->2)]-beta-glucuronopyranoside, were isolated from the aerial parts of Phlomis lunariifolia, in addition to 15 known glycosides. Their structures were elucidated on the basis of extensive 1D and 2D NMR spectroscopic interpretation and chemical degradation.     

Two triacylated and tetraglucosylated anthocyanins from Ipomoea asarifolia flowers

Two triacylated and tetraglucosylated anthocyanins derived from cyanidin were isolated from the flowers of Ipomoea asarifolia and their structures elucidated using chemical, GC, MS and NMR methods (1H and 13C, TOCSY-1D, DQF-COSY, DIFFNOE and HMBC). These complex pigments were found to consist of cyanidin 3-O-[2-O-(6-O-E-caffeoyl-beta-D-glucopyranosyl)]-[6-O-[4-O-(6-O-E-3,5-dihydroxycinnamoyl-beta-D-glucopyranosyl)-E-caffeoyl]-beta-D-glucopyranosyl]-5-O-beta-D-glucopyranoside and cyanidin 3-O-[2-O-(6-O-E-p-coumaroyl-beta-D-glucopyranosyl)]-[6-O-[4-O-(6-O-E-p-coumaroyl-beta-D-glucopyranosyl)-E-caffeoyl]-beta-D-glucopyranosyl]-5-O-beta-D-glucopyranoside.     

Phenolic glycosides from Kaempferia parviflora

Three phenolic glycosides were isolated together with two known flavonol glycosides from the H2O-soluble fraction of rhizomes of Kaempferia parviflora. Their structures were determined to be rel-(5aS,10bS)-5a,10b-dihydro-1,3,5a,9-tetrahydroxy-8-methoxy-6H-benz[b]indeno[1,2-d]furan-6-one 5a-O-[alpha-L-rhamnopyranosyl-(1-->6)-beta-d-glucopyranoside] (1), its rel-5aS,10bR isomer (2), and (2R,3S,4S)-3-O-[alpha-L-rhamnopyranosyl-(1-->6)-beta-d-glucopyranosyl]-3'-O-methyl-ent-epicatechin-(2alpha-->O-->3,4alpha-->4)-(5aS,10bS)-5a,10b-dihydro-1,3,5a,9-tetrahydroxy-8-methoxy-6H-benz[b]indeno[1,2-d]furan-6-one 5a-O-[alpha-L-rhamnopyranosyl-(1-->6)-beta-D-glucopyranoside] (3). The structures were elucidated on the basis of analyses of chemical and spectroscopic evidence.     

Enzymatic synthesis of 2-chloro-4-nitrophenyl 4,6-O-3-ketobutylidene beta-maltopentaoside, a substrate for alpha-amylase.

A transglycosylation reaction with 2-chloro-4-nitrophenyl beta-maltoside as an acceptor was done with 4,6-O-3-ketobutylidene maltopentaose and Bacillus macerans cyclodextrin glucanotransferase in an aqueous solution containing 50% n-propanol, and there were two main transglycosylation products. They were identified as 2-chloro-4-nitrophenyl 4,6-O-3-ketobutylidene beta-maltopentaoside and 2-chloro-4-nitrophenyl 4,6-O-3-ketobutylidene beta-maltohexaoside, and their yields were 30% and 21% respectively on the basis of the decrease of 4,6-O-3-ketobutylidene maltopentaose. For the production of 2-chloro-4-nitrophenyl 4,6-O-3-ketobutylidene beta-maltopentaoside at high substrates concentrations, the addition of n-propanol in this reaction not only increased the solubility of 2-chloro-4-nitrophenyl beta-maltoside sufficiently but also suppressed side reactions.     

Synthesis of modified tetrasaccharide sequences of N-glycoproteins

The tetrasaccharides O-alpha-D-mannopyranosyl-(1----3)-O-[alpha-D- mannopyranosyl-(1----6)]-O-(4-deoxy-beta-D-lyxo-hexopyranosyl)-(1- ---4)-2- acetamido-2-deoxy-alpha, beta-D-glycopyranose (22) and O-alpha-D-mannopyranosyl-(1----3)-O-[alpha-D-mannopyranosyl-(1----6)]-O- beta-D-talopyranosyl-(1----4)-2-acetamido-2-deoxy-alpha, beta-D- glucopyranose (37), closely related to the tetrasaccharide core structure of N-glycoproteins, were synthesized. Starting with 1,6-anhydro-2,3-di-O-isopropylidene-beta-D-mannopyranose, the glycosyl donors 3,6-di-O-acetyl-2-O-benzyl-2,4-dideoxy-alpha-D-lyxo- hexopyranosyl bromide (10) and 3,6-di-O-acetyl-2,4-di-O-benzyl-alpha-D-talopyranosyl bromide (30), were obtained in good yield. Coupling of 10 or 30 with 1,6-anhydro-2-azido-3-O-benzyl-beta-D-glucopyranose to give, respectively, the disaccharides 1,6-anhydro-2-azido-3-O-benzyl-2-deoxy-4-O-(3,6-di-O-acetyl-2-O-benzyl-4 -deoxy- beta-D-lyxo-hexopyranosyl)-beta-D-glucopyranose and 1,6-anhydro-2-azido-3-O-benzyl-2-deoxy-4-O-(3,6-di-O-acetyl-2,4-di-O-ben zyl- beta-D-talopyranosyl)-beta-D-glucopyranose was achieved with good selectivity by catalysis with silver silicate. Simultaneous glycosylation of OH-3' and OH-6' of the respective disaccharides with 2-O-acetyl-3,4,6-tri-O-benzyl-alpha-D-mannopyranosyl chloride yielded tetrasaccharide derivatives, which were deblocked into the desired tetrasaccharides 22 and 37.     

Regioselective phosphorylation of branched cyclodextrins with cyclo-mono-mu-imidotriphosphate

The phosphorylation of the branched cyclodextrins, mono-6-O-(alpha-D-glucopyranosyl)cyclomaltohexaose, mono-6-O-(alpha-D-maltosyl)cyclomaltohexaose, mono-6-O-(alpha-D-glucopyranosyl)cyclomaltoheptaose, and mono-6-O-(alpha-D-maltosyl)cyclomaltoheptaose, in aqueous solution by sodium cyclo-mono-mu-imidotriphosphate (cMITP) was examined. In these reactions, only the 2-OH group of a single alpha-D-glucopyranosyl residue of the cyclodextrin ring was phosphorylated, in a maximum yield of 67%. A possible mechanism for the phosphorylation is discussed.