β-Galactosidase (Aspergillus niger)

Reference code: E-BGLAN
SKU: 700004192

8,000 Units

Content: 8,000 Units
Shipping Temperature: Ambient
Storage Temperature: 2-8oC
Formulation: In 3.2 M ammonium sulphate
Physical Form: Suspension
Stability: > 1 year under recommended storage conditions
Enzyme Activity: β-Galactosidase
EC Number:
CAZy Family: GH35
CAS Number: 9031-11-2
Synonyms: beta-galactosidase; beta-D-galactoside galactohydrolase
Source: Aspergillus niger
Molecular Weight: 125,000
Concentration: Supplied at ~ 4,000 U/mL
Expression: Purified from Aspergillus niger
Specificity: Hydrolysis of terminal non-reducing β-D-galactose residues in β-D-galactosides.
Specific Activity: ~ 140 U/mg (40oC, pH 4.5 on p-nitrophenyl β-D-galactoside)
Unit Definition: One Unit of β-galactosidase activity is defined as the amount of enzyme required to release one µmole of p-nitrophenol (pNP) per minute from p-nitrophenyl-β-D-galactoside (10 mM) in sodium acetate buffer (100 mM), pH 4.5 at 40oC.
Temperature Optima: 60oC
pH Optima: 5
Application examples: Applications established in diagnostics and research within the food and feed, carbohydrate and biofuels industries.

High purity β-Galactosidase (Aspergillus niger) for use in research, biochemical enzyme assays and in vitro diagnostic analysis.

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Certificate of Analysis
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Megazyme publication

A novel enzymatic method for the measurement of lactose in lactose‐free products.

Mangan, D., McCleary, B. V., Culleton, H., Cornaggia, C., Ivory, R., McKie, V. A., Delaney, E. & Kargelis, T. (2018). Journal of the Science of Food and Agriculture, 99, 947-956.

Background: In recent years there has been a surge in the number of commercially available lactose‐free variants of a wide variety of products. This presents an analytical challenge for the measurement of the residual lactose content in the presence of high levels of mono‐, di‐, and oligosaccharides. Results: In the current work, we describe the development of a novel enzymatic low‐lactose determination method termed LOLAC (low lactose), which is based on an optimized glucose removal pre‐treatment step followed by a sequential enzymatic assay that measures residual glucose and lactose in a single cuvette. Sensitivity was improved over existing enzymatic lactose assays through the extension of the typical glucose detection biochemical pathway to amplify the signal response. Selectivity for lactose in the presence of structurally similar oligosaccharides was provided by using a β-galactosidase with much improved selectivity over the analytical industry standards from Aspergillus oryzae and Escherichia coli (EcLacZ), coupled with a ‘creep’ calculation adjustment to account for any overestimation. The resulting enzymatic method was fully characterized in terms of its linear range (2.3-113 mg per 100 g), limit of detection (LOD) (0.13 mg per 100 g), limit of quantification (LOQ) (0.44 mg per 100 g) and reproducibility (≤ 3.2% coefficient of variation (CV)). A range of commercially available lactose‐free samples were analyzed with spiking experiments and excellent recoveries were obtained. Lactose quantitation in lactose‐free infant formula, a particularly challenging matrix, was carried out using the LOLAC method and the results compared favorably with those obtained from a United Kingdom Accreditation Service (UKAS) accredited laboratory employing quantitative high performance anion exchange chromatography - pulsed amperometric detection (HPAEC‐PAD) analysis. Conclusion: The LOLAC assay is the first reported enzymatic method that accurately quantitates lactose in lactose‐free samples.

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Megazyme publication
Measurement of carbohydrates in grain, feed and food.

McCleary, B. V., Charnock, S. J., Rossiter, P. C., O’Shea, M. F., Power, A. M. & Lloyd, R. M. (2006). Journal of the Science of Food and Agriculture, 86(11), 1648-1661.

Procedures for the measurement of starch, starch damage (gelatinised starch), resistant starch and the amylose/amylopectin content of starch, β-glucan, fructan, glucomannan and galactosyl-sucrose oligosaccharides (raffinose, stachyose and verbascose) in plant material, animal feeds and foods are described. Most of these methods have been successfully subjected to interlaboratory evaluation. All methods are based on the use of enzymes either purified by conventional chromatography or produced using molecular biology techniques. Such methods allow specific, accurate and reliable quantification of a particular component. Problems in calculating the actual weight of galactosyl-sucrose oligosaccharides in test samples are discussed in detail.

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Megazyme publication
Measurement of total starch in cereal products by amyloglucosidase-alpha-amylase method: collaborative study.

McCleary, B. V., Gibson, T. S. & Mugford, D. C. (1997). Journal of AOAC International, 80, 571-579.

An American Association of Cereal Chemists/AOAC collaborative study was conducted to evaluate the accuracy and reliability of an enzyme assay kit procedure for measurement of total starch in a range of cereal grains and products. The flour sample is incubated at 95 degrees C with thermostable alpha-amylase to catalyze the hydrolysis of starch to maltodextrins, the pH of the slurry is adjusted, and the slurry is treated with a highly purified amyloglucosidase to quantitatively hydrolyze the dextrins to glucose. Glucose is measured with glucose oxidase-peroxidase reagent. Thirty-two collaborators were sent 16 homogeneous test samples as 8 blind duplicates. These samples included chicken feed pellets, white bread, green peas, high-amylose maize starch, white wheat flour, wheat starch, oat bran, and spaghetti. All samples were analyzed by the standard procedure as detailed above; 4 samples (high-amylose maize starch and wheat starch) were also analyzed by a method that requires the samples to be cooked first in dimethyl sulfoxide (DMSO). Relative standard deviations for repeatability (RSD(r)) ranged from 2.1 to 3.9%, and relative standard deviations for reproducibility (RSD(R)) ranged from 2.9 to 5.7%. The RSD(R) value for high amylose maize starch analyzed by the standard (non-DMSO) procedure was 5.7%; the value was reduced to 2.9% when the DMSO procedure was used, and the determined starch values increased from 86.9 to 97.2%.

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Elucidation of the microstructure of an immuno-stimulatory polysaccharide purified from Korean red ginseng using sequential hydrolysis.

Lee, S. J., In, G., Lee, J. W. & Shin, K. S. (2021). International Journal of Biological Macromolecules, 186, 13-22.

The elucidation of the structural characteristics of polysaccharides from natural sources is generally difficult owing to their structural complexity and heterogeneity. In our previous study, an immuno-stimulatory polysaccharide (RGP-AP-I) was isolated from Korean red ginseng (Panax ginseng C.A. Meyer). The present study aims to elucidate the structural characteristics of RGP-AP-I. Sequential enzyme hydrolysis was performed using four specific glycosylases, and chemical cleavage via β-elimination was carried out to determine the fine structure of RGP-AP-I. The degraded fragments were chemically identified using various chromatographic and spectrometric analyses, including HPLC-UVD, GC–MS, and tandem mass spectrometry. The results indicated that RGP-AP-I comprises a rhamnogalacturonan I (RG-I) backbone with repeating disaccharide units [→2)-Rhap-(1 → 4)-GalAp-(1→] and three side chains substituted at the C(O)4 position of the rhamnose residue in the backbone. The three side chains were identified as a highly branched α-(1 → 5)-arabinan, a branched β-(1 → 4)-galactan, and an arabino-β-3,6-galactan. Our results represent the first findings regarding the fine structure of the immuno-stimulatory polysaccharide RG-AP-I isolated from red ginseng.

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Ancient origin of fucosylated xyloglucan in charophycean green algae.

Mikkelsen, M. D., Harholt, J., Westereng, B., Domozych, D., Fry, S. C., Johansen, I. E., Fangel, J. U., Lęzyk, M., Tao Feng, T., Nancke, L., Mikkelsen, J. D., William G. T. Willats, W. G. T. & Ulvskov , P. (2021). Communications Biology, 4(1), 1-12.

The charophycean green algae (CGA or basal streptophytes) are of particular evolutionary significance because their ancestors gave rise to land plants. One outstanding feature of these algae is that their cell walls exhibit remarkable similarities to those of land plants. Xyloglucan (XyG) is a major structural component of the cell walls of most land plants and was originally thought to be absent in CGA. This study presents evidence that XyG evolved in the CGA. This is based on a) the identification of orthologs of the genetic machinery to produce XyG, b) the identification of XyG in a range of CGA and, c) the structural elucidation of XyG, including uronic acid-containing XyG, in selected CGA. Most notably, XyG fucosylation, a feature considered as a late evolutionary elaboration of the basic XyG structure and orthologs to the corresponding biosynthetic enzymes are shown to be present in Mesotaenium caldariorum.

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RG-I galactan side-chains are involved in the regulation of the water-binding capacity of potato cell walls.

Klaassen, M. T. & Trindade, L. M. (2020). Carbohydrate Polymers, 227, 115353.

Potato cell walls (PCW) are a low value by-product from the potato starch industry. Valorisation of PCW is hindered by its high water-binding capacity (WBC). The composition of polysaccharides and interactions between these entities, play important roles in regulating the WBC in the cell wall matrix. Here, we show that in vivo exo-truncation of RG-I β-(1→4)-D-galactan side-chains decreased the WBC by 6-9%. In contrast, exo-truncation of these side-chains increased the WBC by 13% in vitro. We propose that degradation of RG-I galactan side-chains altered the WBC of PCW, due to cell wall remodelling and loosening that affected the porosity. Our findings reinforce the view that RG-I galactan side-chains play a role in modulating WBC, presumably by affecting polysaccharide architecture (spacing) and interactions in the matrix. Better understanding of structure-function relationships of pectin macromolecules is needed before cell wall by-products may be tailored to render added-value in food and biobased products.

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Rhamnogalacturonan I galactosyltransferase: Detection of enzyme activity and its hyperactivation.

Matsumoto, N., Takenaka, Y., Wachananawat, B., Kajiura, H., Imai, T. & Ishimizu, T. (2019). Plant Physiology and Biochemistry, 142, 173-178.

Rhamnogalacturonan I (RG-I), one of the pectic components of the plant cell wall, is composed of a backbone of repeating disaccharide units of rhamnose and galacturonic acid, and side chains, such as galactans, arabinans, and arabinogalactans. The activity of RG-I galactosyltransferase, which transfers galactosyl residues to rhamnosyl residues in the RG-I backbone, has not been detected until now. Here, we detected galactosyltransferase activity in azuki bean epicotyls using fluorogenic RG-I oligosaccharide acceptors. This enzyme prefers oligosaccharides with a degree of polymerization more than 9. The enzyme activity was detected in the Golgi apparatus, which is the site of pectin synthesis. In vitro hyperactivation of this enzyme was also observed. Moreover, enzyme activity was increased up to 40-fold in the presence of cationic surfactants or polyelectrolytes.

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Mass spectrometry-based identification of carbohydrate anomeric configuration to determine the mechanism of glycoside hydrolases.

Shen, Y. H., Tsai, S. T., Liew, C. Y. & Ni, C. K. (2019). Carbohydrate Research, 476, 53-59.

A rapid mass spectrometry method for determining the anomeric configuration of the sugar at the reducing end of an oligosaccharide was demonstrated. The method was employed to identify the nascent anomeric configuration (i.e., before significant mutarotation occurs) of oligosaccharides released by carbohydrate-active enzymes, which enabled determination of the enzyme mechanism. This method was validated by applying it to various enzymes, including α-glucosidase, β-glucosidases, endoglycoceramidase II, β-galactosidase, and β-amylase.

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Presence of galactose in precultures induces lacS and leads to short lag phase in lactose-grown Lactococcus lactis cultures.

Lorántfy, B., Johanson, A., Faria-Oliveira, F., Franzén, C. J., Mapelli, V. & Olsson, L. (2019). Journal of Industrial Microbiology & Biotechnology, 46(1), 33-43.

Lactose conversion by lactic acid bacteria is of high industrial relevance and consistent starter culture quality is of outmost importance. We observed that Lactococcus lactis using the high-affinity lactose-phosphotransferase system excreted galactose towards the end of the lactose consumption phase. The excreted galactose was re-consumed after lactose depletion. The lacS gene, known to encode a lactose permease with affinity for galactose, a putative galactose–lactose antiporter, was upregulated under the conditions studied. When transferring cells from anaerobic to respiration-permissive conditions, lactose-assimilating strains exhibited a long and non-reproducible lag phase. Through systematic preculture experiments, the presence of galactose in the precultures was correlated to short and reproducible lag phases in respiration-permissive main cultivations. For starter culture production, the presence of galactose during propagation of dairy strains can provide a physiological marker for short culture lag phase in lactose-grown cultures.

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Production of galacto-oligosaccharides from whey permeate using β-galactosidase immobilized on functionalized glass beads.

Eskandarloo, H. & Abbaspourrad, A. (2018). Food Chemistry, In Press.

The conversion of whey permeates to galacto-oligosaccharides (GOS) was studied by the enzymatic action of β-galactosidase from Aspergillus oryzae in a continuous flow packed-bed reactor. A novel method of enzyme immobilization involving covalent immobilization of β-galactosidase on 3-aminopropyl triethoxysilane(3-APTES)-modified glass beads was developed by the cross-linking method. The pH and temperature dependence of the enzymatic efficiency of the glass bead-immobilized enzyme was compared with that of the free enzyme. Increased pH and thermal stabilities were observed for the immobilized enzyme versus the free enzyme. The reusability of the enzyme immobilized packed-bed reactor was studied and only about 4.6% of GOS yield was lost after 8 reuses. Repeated cycle reactions were also carried out to improve the formation of GOS. The results showed that the GOS formation increased and a maximum GOS yield of 39.3% with 56.4% lactose conversion was obtained after the 2nd cycle of passing.

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Safety Information
Symbol : GHS08
Signal Word : Danger
Hazard Statements : H334
Precautionary Statements : P261, P284, P304+P340, P342+P311, P501
Safety Data Sheet
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