Average phase difference theory and 1∶1 phase entrainment in interlimb coordination

The dynamics of coupled biological oscillators can be modeled by averaging the effects of coupling over each oscillatory cycle so that the coupling depends on the phase difference between the two oscillators and not on their specific states. Average phase difference theory claims that mode locking phenomena can be predicted by the average effects of the coupling influences. As a starting point for both empirical and theoretical investigations, Rand et al. (1988) have proposed d/dt= — K sin ), with phase-locked solutions =arcsin( /K), where is the difference between the uncoupled frequencies and K is the coupling strength. Phase-locking was evaluated in three experiments using an interlimb coordination paradigm in which a person oscillates hand-held pendulums. was controlled through length differences in the left and right pendulums. The coupled frequency _c was varied by a metronome, and scaled to the eigenfrequency _v of the coupled system K was assumed to vary inversely with _c. The results indicate that: (1) and K contribute multiplicatively to (2) =0 or = regardless of K when =0; (3) 0 or regardless of when K is large (relative to ); (4) results (1) to (3) hold identically for both in phase and antiphase coordination. The results also indicate that the relevant frequency is _c/_v rather than _c. Discussion high-lighted the significance of confirming =arcsin(/K) for more general treatments of phase-locking, such as circle map dynamics, and for the 11 phase-entrainment which characterizes biological movement systems.     

Molecular cloning and chromosomal localization of a human skeletal muscle PP-1γ1 cDNA

Type-1-protein phosphatase (PP-1) activity is reduced in skeletal muscle from human subjects with insulin resistance (Kida et al. 1990). This reduced phosphatase activity probably leads to the abnormal insulin action for glucose storage observed in insulin-resistant subjects. In the present study, a human homolog of rat liver PP-11 cDNA was isolated from human skeletal muscle. The nucleotide sequence contains a 957-nucleotide open reading frame encoding an amino acid sequence identical to that encoded by rat liver PP-11 cDNA. Northern blot analysis shows PP-11-specific mRNA is expressed in human heart, brain, placenta, lung, liver, skeletal muscle, kidney, and pancreas. PP-11 was localized to human Chromosome 12.     

Evidence of β-carotene 7,8(7′,8′) oxygenase (β-cyclocitral,crocetindial generating) in Microcystis

A -carotene oxygenase is described which occurs in the Cyanobacterium Microcystis. It cleaves -carotene and zeaxanthin specifically at the positions 7,8 and 7,8, while echinenone and myxoxanthophyll are not affected. The oxidative cleavage of -carotene leads to the formation of -cyclocitral and crocetindial and that of zeaxanthin to hydroxy--cyclocitral and crocetindial in nearly stoichiometric amounts. Oxidant is dioxygen as has been demonstrated by high incroporation (86%) of ¹⁸O₂ into -cyclocitral. -Carotene oxygenase is membrane bound, sensitive to sulfhydryl reagents, antioxidants and chelating agents. Iron seems to be an essential part of the enzyme activity. Cofactors necessary for the reaction could not be detected.Abbreviations TLC thin layer-chromatography - PIPES piperazine-N,N-bis-(2-ethanesulfonate) Na - TES 2{[tris-(hydroxymethyl)-methyl]-amino} ethanesulfonic acid Dedicated to Professor G. Drews on occasion of his 60th birthday     

Distribution and Density of Seals in Pil'tun Bay (Northeastern Sakhalin) in the Summer–Autumn Period

The results of seal counts performed during the summer–autumn period of the years 1999–2001 in Pil'tun Bay from onboard Zodiak motor boats and Mi-8 and Mi-8MTV helicopters have been analyzed. The information is presented on the density of seals in the bay. The heterogeneous distribution pattern of seals in the bay and the presence of intermittent migrations provide favorable conditions for foraging in young animals.     

Ganglioside-cholera toxin interactions: a binding and lipid monolayer study

Summary On t.l.c. plates ¹²⁵I-cholera toxin binds to a disialoganglioside tentatively identified as GD_lb with about 10 times less capacity than to ganglioside GM₁. Binding of labeled toxin to both gangliosides was abolished in presence of excess amounts of unlabeled B subunit. Ganglioside extracts from human or pig intestinal mucosa showed toxin binding to gangliosides GM₁ and GD_1b. In ganglioside-containing lipid monolayers the penetration of the toxin was independent of the ganglioside binding capacity.Abbreviations GM₂ Gal-NAc14Gal(3-2NeuAc)14G1c1Cer - GM₁ Gal3Ga1-NAc14Gal(32NeuAc)14G1c11Cer - GD_1a NeuAc23Ga113Gal-NAc14Gal(32NeuAc)14G1c11Cer - GD1b Gall3Gal-NAcl4Gal(32NeuAc82NeuAc)14Glc11Cer - GT_1b NeuAc23Ga113Ga1-NAcal4Gal(3-2NeuAc82NeuAc)14G1c11Cer - dpPC 1,2-hexadecanoyl-sn-glycero-3-phosphocholine - dpPE 1,2-hexadecanoyl-sn-glycero-3-phosphoethanolamine     

Metal Cations Defibrillize the Amyloid Beta-Protein Fibrils

Amyloid beta-protein (A) is the major constituent of amyloid fibrils composing -amyloid plaques and cerebrovascular amyloid in Alzheimer's disease (AD). We studied the effect of metal cations on preformed fibrils of synthetic A by Thioflavin T (ThT) fluorescence spectroscopy and electronmicroscopy (EM) in negative staining. The amount of cross beta-pleated sheet structure of A 1–40 fibrils was found to decrease by metal cations in a concentration-dependent manner as measured by ThT fluorescence spectroscopy. The order of defibrillization of A 1–40 fibrils by metal cations was: Ca²⁺ and Zn²⁺ (IC₅₀ = 100 M) > Mg²⁺ (IC₅₀ = 300 M) > Al³⁺ (IC₅₀ =1.1 mM). EM analysis in negative staining showed that A 1–40 fibrils in the absence of cations were organized in a fine network with a little or no amorphous material. The addition of Ca²⁺, Mg²⁺, and Zn²⁺ to preformed A 1–40 fibrils defibrillized the fibrils or converted them into short rods or to amorphous material. Al³⁺ was less effective, and reduced the fibril network by about 80 % of that in the absence of any metal cation. Studies with A 1–42 showed that this peptide forms more dense network of fibrils as compared to A 1–40. Both ThT fluorescence spectroscopy and EM showed that similar to A 1–40, A 1–42 fibrils are also defibrillized in the presence of millimolar concentrations of Ca²⁺. These studies suggest that metal cations can defibrillize the fibrils of synthetic A.     

The polypeptide structure and assembly of Ly-2/3 heterodimers

Mild reduction of mature, thymic Ly-2/3 heterodimers of M _r 67 000 resulted in dissociation into three individual polypeptide chains, , , and , of respective M _r values 38000, 35000, and 30000. The and chains were both immunoprecipitated by a monoclonal antibody directed to the Ly-2.1 epitope whereas the Ly-3.1 antibody bound only the chain. The possibility that the and chains of each heterodimer established their interchain links within a labile precursor protein in which a and segments were fused was considered but discounted by the finding that in mice heterozygous for both Ly-2 and Ly-3 loci, the Ly-2 product of one chromosome was not exclusively joined to Ly-3 structures coded by the same chromosome. By utilizing ionic detergents which selectively alter the charge of intrinsic membrane proteins, both Ly-2 and Ly-3 polypeptides were shown to have membrane insertion sites. It is suggested that as a consequence of their likely synthesis on membrane-bound polysomes, newly synthesized Ly-2 and Ly-3 structures accumulate within the same subcellular compartment — the membranes of the rough endoplasmic reticulum. Their elevated concentration within this space may facilitate a low affinity binding interaction between Ly-2 and Ly-3 which is later stabilized by interchain disulfide bond formation.Abbreviations used in this paper BSA bovine serum albumin - DOC sodium deoxycholate - DTT dithiothreitol - HA hemagglutinin - HTAB hexadecyltrimethylammonium bromide - RER rough endoplasmic reticulum - SDS-PAGE sodium dodecyl sulfate polyacrylamide gel electrophoresis - TX100 Triton X-100     

The use of BRM-activated killer cells in adoptive immunotherapy: A pilot study with nine advanced cancer patients

Adoptive immunotherapy using MHC-nonrestricted-lymphocytes, peripheral blood T cells and NK cells was devised. Peripheral blood mononuclear cells (3 x 107) were selected by immobilization to anti-CD3 monoclonal antibody for 4 days and cultured for 2 weeks in the presence of IL-2. Thereafter they were reactivated by 500 U/ml of IFN- and 1000 U/ml of IL-2 for 1 hour. Enhancement of NK and LAK activities was confirmed. Peripheral blood T cells proliferated in response to immobilized anti-CD3 antibody (3% to 30%). Approximately 6 x 109 BRM-activated killer (BAK) cells composed of CD56+ T cells and CD56+ NK cells, were dispensed to cancer patients via intravenous drip infusion. Nine patients were treated with BAK cells every 2 weeks or every month on an outpatient basis. During the course of adoptive immunotherapy, the crossed affinity immunoelectrophoresis (CAIE) pattern of serum immunosuppressive acidic protein (IAP) was analysed. Both the production and glycosylation pattern of IAP is changed in response to tumor enlargement and may therefore act as a marker of the disease progression. During the course of BAK therapy, the glycosylation IAP pattern of 6 patients changed from tumor (T) to normal (N). In addition, the performance status of all patients was maintained at 90–100% of the Karnofsky scale and any side effects including fever were not observed during treatments with BAK cells. Moreover, the overall quality of life (QOL) of the patients, scored at the Face scale was favorable. In addition, blood levels of activated T cells producing IFN- were assayed as an indication marker of BAK therapy. The normal range of IFN- producing T cells comprised 6.9 ± 0.9% of peripheral blood mononuclear cells (PBMC), according to a single cell FACScan analyses of PBMCs derived from normal individuals. IFN- producing T cells of Patients No. 8 and 9, who received extensive chemotherapy before initiation of BAK therapy, comprised only 0.2% and 2% of PBMC, respectively. These patients died 3 and 6 months after beginning BAK therapy. Peripheral blood T cells of Patients Nos. 1–7 proliferated in response to immobilized anti-CD3 antibody and the frequency of IFN- producing T cells in PBMC preparation of these patients were over 3% before initiation of BAK therapy. Since our data show a positive correlation between survival time and initial T cell counts, a low frequency of these cells may contraindicate BAK therapy.     

Concluding remarks

On the basis of symposium contributions onChlorella, Hibbertia, Eucalyptus, Ambrosia and on numerical approaches some fundamental problems of (bio)systematics, evolution, and taxonomic categories are discussed: Methods available for analysing affinities; conflicting evidence from phenetic, biochemical, cytogenetic and other analyses; further classification problems in cases of intermediacy, etc. While sibs of various levels and their natural hierarchy often can be objectively defined, this appears impossible for particular taxonomic levels itself (e. g. species). A single objective taxonomic system of organisms is unrealistic. Certain guiding lines for relative and practicable concepts of species and genus are proposed.Presented at the symposium Speciation and the Species Concept during the XIIth International Botanical Congress, Leningrad, July 8, 1975.     

Developmental aspects of long-term callus culture ofVicia faba L.

Summary Vicia faba callus line (VFS 1), isolated from expiants of immature embryo, grew satisfactorily onMurashige andSkoog complete medium with 1.38 M 2,4-D, or with 0.92 M 2,4-D to which 1.0 M kinetin was added. It also grew well on the B 5 modified medium containing 2.3 M 2,4-D and 25.0 M kinetin. On the last of these media the cultures grew more uniformly and without necrosis. They also showed diminishing variation in polyploidy in favour of diploids and corresponding aneuploids (hypodiploids).After being cultured for nearly three years on MS containing 1.38 M 2,4-D, 8–33% of cultures of VFS 1 were able to regenerate roots when transferred to either MS half strength with 5.37 M NAA, or to a medium without 2,4-D, or else to media with the addition of kinetin only (in various concentrations).     

An elongated segment of DNA observed between two human α globin genes

Summary Detailed restriction enzyme analysis of the DNA from a Chinese female showed that one of her chromosomes had a >17.5 kb deletion of DNA, including the , 2, and 1 globin genes, which is present in many Southeast Asians with an -thalassemia-1 chromosome. Her normal chromosome had the expected cluster of -like globin genes (5----2-1-3), but the segment of DNA between the two globin genes was elongated by some 0.5–0.7 kb. Analyses of various restriction sites suggested that this normal variant of the human globin gene complex is due to a crossover between a normal chromosome with () and a chromosome with an -thalassemia-2 (–^3.7) and an -21-hybrid gene.     

Relative positions of homologous chromosomes or groups in male and female metaphase figures

Summary This study investigates statistically, with computer assistance, the square distances between chromosome centromeres in homologous pairs or groups after circularizing transformation. By the method described here, it is possible to eleminate all subjective measurments and just use the coordinates x _i and y _i of the centromeres. The values obtained for homologous chromosomes are shown to be specific but not always small. Low square distance values occur in greater number for the chromosomes that are most frequently involved in aneuploïdies. This is true for acrocentric chromosomes which, moreover, tend to lie close together significantly more often in female than in male mitoses; it is also true for group 17–18 especially in males, and of XX in female mitoses. Furthermore, we find significantly low square distances in chromosome pair 1.     

The mitochondrial membrane potential (deltapsi(m)) in apoptosis; an update

Mitochondrial dysfunction has been shown to participate in the induction of apoptosis and has even been suggested to be central to the apoptotic pathway. Indeed, opening of the mitochondrial permeability transition pore has been demonstrated to induce depolarization of the transmembrane potential (_m), release of apoptogenic factors and loss of oxidative phosphorylation. In some apoptotic systems, loss of _m may be an early event in the apoptotic process. However, there are emerging data suggesting that, depending on the model of apoptosis, the loss of _m may not be an early requirement for apoptosis, but on the contrary may be a consequence of the apoptotic-signaling pathway. Furthermore, to add to these conflicting data, loss of _m has been demonstrated to not be required for cytochrome c release, whereas release of apoptosis inducing factor AIF is dependent upon disruption of _m early in the apoptotic pathway. Together, the existing literature suggests that depending on the cell system under investigation and the apoptotic stimuli used, dissipation of _m may or may not be an early event in the apoptotic pathway. Discrepancies in this area of apoptosis research may be attributed to the fluorochromes used to detect _m. Differential degrees of sensitivity of these fluorochromes exist, and there are also important factors that contribute to their ability to accurately discriminate changes in _m.     

Convergent synthesis of neoglycopeptides by coupling of 2-bromoethyl glycosides to cysteine and homocysteine residues in T cell stimulating peptides

The 2-bromoethyl -glycosides of the disaccharide galabiose [Gal(1-4)Gal] and the trisaccharides globotriose [Gal(1-4)Gal(1-4)Glc] and 3-sialyllactose [Neu5Ac(2-3)Gal(1-4)Glc] have been prepared by improved routes. The 2-bromoethyl glycosides were then used in cesium carbonate promoted alkylations of the sulfhydryl groups of cysteine and homocysteine residues in T cell stimulating peptides. This convergent and general approach was used to prepare 16 neoglycopeptides which were obtained in 52–95% yields after purification by HPLC. 1H NMR spectroscopy revealed that -elimination and epimerization of neoglycopeptide stereocentres did not occur during the synthesis.     

Immunohistological analyses of neutral glycosphingolipids and gangliosides in normal mouse skeletal muscle and in mice with neuromuscular diseases

The expression of neutral glycosphingolipids (GSLs) and gangliosides was investigated in cryosections of normal mouse skeletal muscle and in muscle of mice with neuromuscular diseases using indirect immunofluorescence microscopy. Transversal and longitudinal sections were immunostained with specific polyclonal antibodies against lactosylceramide, lacto-N-neotetraosylceramide, globoside, G_M3(Neu5Ac), G_M3(Neu5Gc) and G_M1(Neu5Ac) as well as monoclonal anti-Forssman GSL antibody. In normal CBA/J mouse muscle (control) the main immunohistochemically detected ganglioside was G_M3(Neu5Ac) followed by moderately expressed G_M3(Neu5Gc) and G_M1. The neutral GSLs lactosylceramide and globoside were stained with almost identical, high fluorescence intensity. Low amounts of lacto-N-neotetraosylceramide and trace quantities of Forssman GSL were immunostained. All GSLs were detected in the sarcolemma, but also in considerable amounts at the intracellular level. Mice with neuromuscular diseases were the A2G-adr mouse mutant (a model for human recessive myotonia of Becker type), the BL6-wr mutant (a model for motor neuron disease) and the BL10-mdx mouse mutant (a model for human Duchenne muscular dystrophy). No changes in GSL expression were found in the A2G-adr mouse, while muscle of the BL6-wr mouse showed increased intensity of immunofluorescence in stainings with anti-lactosylceramide and anti-G_M3(Neu5Ac) antibodies. Muscle of BL10-mdx mice showed the most prominent changes in GSL expression with reduced fluorescence intensity for all antibodies. Major differences were not observed in the intensities of GSLs, but there were significant differences in the patterns of distribution on plasma membrane and at the subcellular level. The exact nature and pathogenesis of these changes should be elucidated since such investigations could furnish advances in understanding the functional role of neutral GSLs and gangliosides in normal as well as in diseased muscle. Abbreviations: BSA, bovine serum albumin; DAPI, 4, 6-diamidine-2-phenylindole-dihydrochloride; DTAF, dichlorotriazinylamino-fluorescein; GSL(s), glycosphingolipid(s); Neu5Ac,N-acetylneuraminic acid; Neu5Gc,N-glycolylneuraminic acid [53]; PBS, phosphate buffered saline. The designation of the following glycosphingolipids follows the IUPAC-IUB recommendations [54] and the nomenclature of Svennerholm [55]. Lactosylceramide or LacCer, Gal1-4Glc1-1Cer; gangliotriaosylceramide or GgOse₃Cer, GalNAc1-4Gal1-4Glc1-1Cer; globotriaosylceramide or GbOse₃Cer, Gal1-4Gal1-4Glc1-1Cer; gangliotetraosylceramide or GgOse₃Cer, Gal1-3GalNAc1-4Gal1-4Glc1-1Cer; globotetraosylceramide or GbOse₄Cer, GalNAc1-3Gal1-4Gal1-4Glc1-1Cer; lacto-N-neotetraosylceramide or nLcOse₄Cer, Gal1-4GlcNAc1-3Gal1-4Glc1-1Cer; Forssman GSL or GbOse₅Cer, GalNAc1-3GalNAc1-3GAl1-4Gal1-4Glc1-1Cer; G_M3, II³Neu5Ac-LacCer; G_M1, II³Neu5Ac-GgOse₄Cer.     

Mode of uptake of sterol by Arthrobacter simplex

The mechanism of uptake of water-insoluble -sitosterol by a newly isolated strain of Arthrobacter simplex SS-7 was studied. The production of an extracellular sterol-pseudosolubilizing protein during growth of A. simplex on -sitosterol was demonstrated by isolating the factor from the cell-free supernatant and its subsequent purification by Sephadex G-150 column chromatography. The M _r of the purified sterol-pseudosolubilizing protein determined by SDS–PAGE was 19kDa. The rate of sterol pseudosolubilization (5.2×10^–3g l^–1h^–1) could not adequately account for the rate of sterol uptake (72×10^–3g l^–1h^–1) and the specific growth rate (56×10^–3 h^–1). However in the unfavourable growth condition, when the cells were treated with sodium azide at the level of 30–60% of MIC, the sterol pseudosolubilization accounted for nearly 74% of the total growth containing 96% free cells. Cellular adherence to substrate particles was found to play an active role in the normal growth of the strain on -sitosterol. Unlike sodium acetate-grown cells, whose surface activity was negligible (60mNm^–1), the sterol-grown cells had strong surface activity (40mNm^–1). The high lipid content and long chain fatty acids in the cell-wall of -sitosterol-grown cells probably contribute to the high sterol adherence activity of the cells.     

Fever and feeding: Differential actions of macrophage inflammatory protein-1 (MIP-1), MIP-1α and MIP-1β on rat hypothalamus

Changes in body temperature (T_b) and feeding were characterized in unrestrained rats following the micro-injection into the anterior hypothalamic preoptic area (AH/POA) of macrophage inflammatory protein-1 (MIP-1), MIP-1 or MIP-1. After the rats recovered from the stereotaxic implantation of a single guide tube placed in the AH/POA, either one of the MIP-1 compounds or control CSF was micro-injected in a volume of 1.0 l into this area. Changes in body temperature (T_b) and food and water intakes were monitored throughout each experiment. When micro-injected into the AH/POA in a dose of 28 or 280 pg, doublet MIP-1 and MIP-1 evoked a monophasic fever which increased above baseline to a mean maximum of 2.17±0.14°C and 2.1±0.24°C, respectively. MIP-1 micro-injected similarly evoked a biphasic fever, with the T_b declining transiently at the 30 min point 0.4°C lower than the congruent rises in T_b evoked by doublet MIP-1 or MIP-1. The secondary rise in T_b induced by MIP-1 had a latency of 1.5–2.0 hrs and reached a maximum of 1.56±0.16°C. Although all three cytokines significantly attentuated the rats' mean intake of food during the 24 hr interval after their micro-injection into the AH/POA, doublet MIP-1 exerted the most potent anorexic effect in comparison to that of the saline control rats. However, neither body weight nor intake of water was altered significantly by the three cytokines. These results demonstrate a functional distinction between the febrile actions of MIP-1 and MIP-1 on cells of the AH/POA. It is envisaged that MIP-1 underlies the initial phase of a pyrogen-induced fever, whereas MIP-1 could mediate the secondary phase of a biphasic fever. Clearly, the localized elevation of either MIP-1 or MIP-1 in the AH/POA perturbs the neuronal mechanism underlying the set-point for body temperature. Thus, both MIP-1 and MIP-1 could play a functional role in the pathological events occurring in neurons of the AH/POA in response to an endotoxin or other pyrogen challenge.     

Monoclonal antibodies that recognize the trisaccharide epitope Galα1-3Galβ1-4GlcNAc present on Ehrlich tumor cell membrane glycoproteins

Monoclonal antibodies were prepared against the trisaccharide Gal1-3Gal1-4GlcNAc, a sequence which occurs on the surface of Ehrlich ascites tumor cells as well as in thyroglobulin, laminin and a variety of other proteins. This was accomplished by immunizing BALB/c mice with the fraction of Ehrlich cell membrane glycoproteins obtained by affinity chromatography on aGriffonia simplicifolia I (GS I) column which selectively binds -d-galactosyl-terminated structures. Detection of Gal1-3Gal1-4GlcNAc-specific antibodies was accomplished by employing glycoproteins containing the trisaccharide sequence; fusion with spleen cells from an immunized mouse was accomplished in the presence of polyethylene glycol (PEG1500). An enzyme-linked immunosorbent assay (ELISA) system was used to identify two clones (2.10G and 6.8E), which recognized the desired trisaccharide conjugate. These clones also recognized a thyroglobulin fraction isolated by GS I affinity chromatography and murine laminin, both of which possess the Gal1-3Gal1-4GlcNAc sequence. Inhibition of antibody-trisaccharide reactivity, examined employing an ELISA assay, revealed that two trisaccharides, Gal1-3Gal1-4GlcNAc/Glc, were the best inhibitory haptens; Gal1-4GlcNAc (LacNAc), Gal1-3Gal and Gal1-4Glc (lactose) were poor inhibitors. Indirect immunofluorescence staining of unfixed Ehrlich cells using the monoclonal antibody at 4° C revealed fluorescence over the entire cell surface. Indirect immunogold labeling of semithin and ultrathin sections of aldehyde fixed and Lowicryl K4M-embedded Ehrlich cells resulted in specific labeling of the cell surface and internal structure. Immunoblot analysis revealed that removal of the -galactosyl residues of laminin by -galactosidase abolished reactivity with the monoclonal antibodies. The availability of this antibody, which belongs to the IgM family of immunoglobulins, now makes possible the detection of this sugar sequence on cells and tissue sections, as well as on glycoproteins in solution.     

Plant regeneration from cytoplasmic hybrids of rice (Oryza sativa L.)

Summary We obtained cybrid plants by electrofusing -irradiated protoplasts of a cytoplasmic male-sterile line A-58 CMS (Oryza sativa L.) and iodoacetamide (IOA)-treated protoplasts of the fertile (normal) rice cultivar Fujiminori. The cybridity of the plants was confirmed by mitochondrial (mt) DNA restriction endonuclease, and plasmid-like DNA analyses, and by isozyme, cytological and morphological investigations. The chromosome number of the cybrid plants is 24.     

Enzymic activities and galactomannan mobilisation in germinating seeds of fenugreek (Trigonella fœnum-graecum L. Leguminosae)

Summary The activities of -galactosidase, -mannosidase and -mannosidase were determined in extracts from the endosperm and from the embryo of fenugreek seeds at different stages of germination. Endosperm homogenates contained little or no activity of the above enzymes in the early stages of germination, before the reserve galactomannan began to be mobilised. The onset of galactomannan breakdown coincided with the appearance of -galactosidase and -mannosidase activities, which increased throughout the period of galactomannan degradation and then remained constant. A similar rise in -galactosidase and -mannosidase activities occurred during galactomannan breakdown in dry-isolated endosperms incubated under germination conditions. The increase could be suppressed by metabolic inhibitors which also inhibit galactomannan breakdown. Embryo homogenates contained high -galactosidase, high -mannosidase and some -mannosidase activity at all stages of germination.No oligomannosyl -1,4 phosphorylase activity could be detected either in the endosperm or in the embryo.It is concluded that the galactomannan of fenugreek is broken down by a series of hydrolases secreted by the aleurone layer of the endosperm. They include -galactosidase, -mannosidase and probably also endo--mannanase.This is part four in a series of papers dealing with galactomannan metabolism. Part three: Planta (Berl.) 106, 44–60 (1972).