L-天门冬素[天冬酰胺酸]/L-谷氨酸/氨检测试剂盒 L-Asparagine/L-Glutamine/ Ammonia 货号:K-ASNAM Megazyme中文站

L-天门冬素[天冬酰胺酸]/L-谷氨酸/氨检测试剂盒

英文名:L-Asparagine/L-Glutamine/ Ammonia

货号:K-ASNAM

规格:100 Assays (50 of each)

分析物意义:  炸、烘、烤土豆或其他食品中的丙烯酰胺的前提

Megazyme检测试剂盒优点:产品新颖,不到20分钟可以检测3种组分。可用于手工或酶标仪自动检测。

 

The L-Asparagine/L-Glutamine/Ammonia test kit is a novel method for the specific, convenient, cost effective and rapid measurement and analysis of L-asparagine, L-glutamine and ammonia as acrylamide precursors in the food industry, or as cell culture media/supernatant components, or in other materials.

Extended cofactors stability. Dissolved cofactors stable for > 1 year at 4oC.

UV-method for the determination of L-Asparagine, L-Glutamine 
and Ammonia in potatoes, foodstuffs and cell culture media

Principle:
                               (glutaminase)
(1) L-Glutamine + H2O → L-glutamate + NH4+

                             (microbial glutamate dehydrogenase)
(2) NH4+ + 2-oxoglutarate + NADPH → L-glutamate + NADP+ + H2O

                                (asparaginase)
(3) L-Asparagine + H2O → L-aspartate + NH4+

Kit size:                              50 assays of each
Method:                              Spectrophotometric at 340 nm
Reaction time:                   ~ 20 min
Detection limit:                  0.50 mg/L (L-asparagine)
                                           0.54 mg/L (L-glutamine)
                                           0.06 mg/L (ammonia)
Application examples:
Potatoes, potato products, vegetables, cereals and other materials
(e.g. biological cultures, samples, etc.)
Method recognition:          Novel method

Advantages

  • Very rapid reaction due to use of uninhibited glutamate dehydrogenase
     
  • All enzymes supplied as stabilised suspensions
     
  • Only kit available
     
  • Very cost effective
     
  • All reagents stable for > 2 years after preparation
     
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
     
  • Standard included

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乙醛检测试剂盒 Acetaldehyde Assay Kit 货号:K-ACHYD Megazyme中文站

乙醛检测试剂盒

英文名:Acetaldehyde Assay Kit

货号:K-ACHYD

规格:50 assays (manual) / 500 assays

紫外吸光度法测定食品、饮料和其他原料中的乙醛。

原理:
                                     (醛脱氢酶)
Acetaldehyde + NAD+ + H2O → acetic acid + NADH + H+
试剂盒规格: 50次检测
方法:分光光度计,340nm
反应时间: ~4min
检测限制: 0.18 mg/L
适用样品:葡萄酒、香槟酒、啤酒、烈酒、白兰地、乳制品(如酸奶)面包、果汁、软饮料、可可粉、蔬菜和水果制品、咖啡和其他原料(如:生物培养基,样品等。)
方法认证: 该实验方法已通过MEBAK和瑞士认证。

优点:
不会浪费醛脱氢酶溶液(提供稳定的悬浮液)
价格低廉(每次检测成本)
所有试剂配置后的稳定性>2年
操作简单
网站提供Mega-Calc计算软件,处理原始数据更方便
包含标准品

 

Advantages

  • No wasted aldehyde dehydrogenase solution (stable suspension supplied)
  • Very competitive price (cost per test)
  • All reagents stable for > 2 years after preparation
  • Simple format
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
  • Standard included
  • Extended cofactors stability
  • Suitable for manual, microplate and auto-analyser formats

UV-method for the determination of Acetaldehyde in foodstuffs, beverages and other materials

Principle:
(aldehyde dehydrogenase)
(1) Acetaldehyde + NAD+ + H2O → acetic acid + NADH + H+

Kit size: 50 assays (manual) / 500 (microplate)
/ 500 (auto-analyser)
Method: Spectrophotometric at 340 nm
Reaction time: ~ 4 min
Detection limit: 0.18 mg/L
Application examples:
Wine, champagne, beer, liqueurs, brandy, dairy products (e.g. yogurt),
bread, fruit juices, soft drinks, cocoa, vegetable and fruit products,
coffee, and other materials (e.g. biological cultures, samples, etc.)
Method recognition:
Methods based on this principle have been accepted by MEBAK

 

Q1. Sometimes a negative absorbance change is obtained for the blank samples, is this normal? Should the real value (negative absorbance change) or “0” be used in the calculation of results?

Sometimes the addition of the last assay component can cause a small negative absorbance change in the blank samples due to a dilution effect, and in such cases it is recommended that the real absorbance values be used in the calculation of results.

Q2. Should the pH of the sample be adjusted even for samples in acidic media?

The pH of the assay solution after the sample is added should be the same as that of the assay buffer that is supplied with the kit.
Low sample volumes (e.g. 0.1 mL) are not likely to affect the pH of the assay solution and therefore may not require pH adjustment.
Samples above 0.1 mL are more likely to affect the pH of the assay solution and therefore the pH of these samples should be adjusted as described in the data booklet, prior to addition to the assay.

Q3. There is an issue with the performance of the kit; the results are not as expected.

If you suspect that the Megazyme test kit is not performing as expected such that expected results are not obtained please do the following:

  1. Ensure that you have tested the standard sample that is supplied with the Megazyme test kit.
  2. Send the results of the kit standard, blank samples and the results obtained for your sample,  in the relevant MegaCalc spreadsheet (if available) to Megazyme (cs@megazyme.com). Where available the relevant MegaCalc spreadsheet can be downloaded from where the product appears on the Megazyme website.
  3. State the kit lot number being used (this is found on the outside of the kit box).
  4. State which assay format was used (refer to the relevant page in the kit booklet if necessary).
  5. State exact details of any modifications to the standard procedure that is provided by Megazyme.
  6. State the sample type and describe the sample preparation steps if applicable.

Q4. Can perchloric acid be used to deproteinise/clarify samples prior to analysis using the Acetaldehyde Assay Kit (K-ACHYD)? If so, how should such an extraction be performed?

Yes.  Perchloric acid extraction can be used in conjunction with this kit, and should be performed as follows:
WARNING:  If you have not worked with perchloric acid before, you must consult your safety officer for advice.  Also, depending on the nature of the samples, it may be possible to reduce the concentration of perchloric acid, to for example 0.3 M (i.e. in the case of plasma).  It is thus very important to determine if this is possible for each type of sample used, in order to reduce the risk from working with concentrated perchloric acid.

Liquid samples:

  1. Carefully add an equal volume of ice cold 3 M perchloric acid and homogenise/fully disperse the sample (as appropriate).
  2. After 15 min incubation on ice (or in a refrigerator), centrifuge at 3,000 x g for 15 min at 4oC.
  3. Neutralise by the slow addition of 2 M KOH.
  4. Incubate on ice (or in a refrigerator) until the potassium perchlorate has settled out by gravity (approximately 10 min), and then simply remove some of the clear supernatant and use directly in the assay.  


Solid samples:

  1. Accurately weigh approx. 5 g of homogenised sample into a beaker containing 20 mL of 1 M perchloric acid and very carefully homogenise with an Ultraturrax® (or equivalent) for 5 min.
  2. Carefully add approx. 40 mL of distilled water and neutralise using 2 M KOH (using pH test strips for example).  Quantitatively transfer the contents to a 100 mL volumetric flask and fill to the mark with distilled water.  If a fat layer develops, make sure this is above the mark, and the aqueous layer is at the mark.
  3. Incubate in a refrigerator for approx. 20 min to allow separation of fat and precipitation of potassium perchlorate.
  4. Filter through Whatman No. 1 filter paper, discarding the first few mL of filtrate, and use directly in the assay.

Q5. The pH of my sample is low (pH ~ 3.0), do I need to adjust this before I use the sample in the kit assay?

The final pH of the kit assay after the sample is added should not change from what it should be (as stated in the kit for the assay buffer). If it does change then the sample will require pH adjustment. In most cases the sample volume being used is low relative to the final assay volume and in this case the pH of the kit assay is unlikely to be affected.

Q6. How can I work out how much sample to extract and what dilution of my sample should be used in the kit assay?

Where the amount of analyte in a liquid sample is unknown, it is recommended that a range of sample dilutions are prepared with the aim of obtaining an absorbance change in the assay that is within the linear range.
Where solid samples are analysed, the weight of sample per volume of water used for sample extraction/preparation can be altered to suit, as can the dilution of the extracted sample prior to the addition of the assay, as per liquid samples.

Q7. Can you explain, step by step, how to follow the method and perform the kit assay?

For users who are not familiar with how to use the Megazyme tests kits then it is recommended that they follow this example, e.g. D-Fructose/D-Glucose Assay kit K-FRUGL (http://secure.megazyme.com/D-Fructose-D-Glucose-Assay-Kit):

1. The kit components are listed on pages 2-3 of the kit booklet.
2. Prepare the kit reagents as described on page 3.
3. For separate measurements of glucose and fructose follow procedure A on page 4.
4. Pipette the volumes listed for water, sample, solution 1 and solution 2 into 3 mL, 1 cm pathlength cuvettes. Duplicate sample assays and duplicate blanks are recommended. Mix the contents of each cuvette by inversion (seal the cuvette using parafilm or a plastic cuvette cap – do not use a finger) then after ~3 min record the first absorbance reading of each cuvette at 340 nm (this is reading A1).
5. Then add suspension 3 and mix the contents of each cuvette by inversion. Incubate for 5 minutes then record the absorbance reading of each cuvette at 340 nm (this is reading A2). NB. It is essential that the reaction is compete. To assess this, record the absorbances at ~ 2 minute intervals and until the absorbance plateaus. A stable absorbance indicates that the reaction is complete. If the absorbance continues to increase then continue to record absorbances until it plateaus and only then record absorbance reading A2.
6. Then add suspension 4 and mix the contents of each cuvette by inversion. Incubate for 5 minutes then take absorbance reading of each cuvette at 340 nm (this is reading A3). NB. As above, assess that the reaction has completed by take subsequent readings at ~2 min intervals.
7. For simple, automated results analysis, input the absorbance readings (A1, A2, A3) for samples and blanks into the K-FRUGL MegaCalc.

To ensure that the assay is working, and being performed correctly it is recommend that the test is performed using the standard sample that is provided with the kit and to obtain the expected values before proceeding to test real samples.
It is recommend that new users also watch this video which highlights how to perform the assays.
Many of the other Megazyme test kits follow a similar format.

Q8. I have some doubts about the appearance/quality of a kit component what should be done?

If there are any concerns with any kit components, the first thing to do is to test the standard sample (control sample) that is supplied with the kit and ensure that the expected value (within the accepted variation) is obtained before testing any precious samples. This must be done using the procedure provided in the kit booklet without any modifications to the procedure. If there are still doubts about the results using the standard sample in the kit then send example results in the MegaCalc spread sheet to your product supplier (Megazyme or your local Megazyme distributor).

Q9. How much sample should be used for the clarification/extraction of my sample?

The volume/weight of sample and total volume of the extract can be modified to suit the sample. This will ultimately be dictated by the amount of analyte of interest in the sample and may require empirical determination. For low levels of analyte the sample:extract volume ratio can be increased (i.e. increase the sample and/or decrease the total extraction volume).

Alternatively, for samples with low concentrations of analyte, a larger sample volume can be added to the kit assay. When altering the sample volume adjust the distilled water volume added to the assay accordingly so that the total assay volume is not altered.

Q10. Can the manual assay format be scaled down to a 96-well microplate format?

The majority of the Megazyme test kits are developed to work in cuvettes using the manual assay format, however the assay can be converted for use in a 96-well microplate format. To do this the assay volumes for the manual cuvette format are reduced by 10-fold. The calculation of results for the manual assay format uses a 1 cm path-length, however the path-length in the microplate is not 1 cm and therefore the MegaCalc spreadsheet or the calculation provided in the kit booklet for the manual format cannot be used for the micropalate format unless the microplate reader being used can.

There a 3 main methods for calculation of results using the microplate format:

  1. The easiest method is to use a microplate reader that has a path-length conversion capability (i.e. the microplater reader can detect the path-length of each well and convert the individual readings to a 1 cm path-length). This will allow values to be calculated using the MegaCalc calculation software which can be found where the product is located on the Megazyme website.
  2. Perform a standard curve of the analyte on each microplate that contains test samples and calculate the result of the test samples from the calibration curve (concentration of analyte versus absorbance).
  3. Perform a standard curve of the analyte in both the cuvette format (i.e. with a 1 cm path-length) and the 96-well microplate format and use these results to obtain a mean conversion factor between the cuvette values and the microplate values. Subsequent assays in the microplate format can then be converted from the calculated conversion factor.

Q11. Can the test kit be used to measure biological fluids and what sample preparation method should be used?

The kit assay may work for biological fluids assuming that inositol is present above the limit of detection for the kit after any sample preparation (if required). Centrifugation of the samples and use of the supernatant directly in the kit assay (with appropriate dilution in distilled water) may be sufficient. However, if required a more stringent sample preparation method may be required and examples are provided at the following link:http://www.megazyme.com/docs/analytical-applications-downloads/biological_samples_111109.pdf?sfvrsn=2

The test kit has not been tested using biological fluids as samples because it is not marketed or registered as a medical device. This will therefore require your own validation.

Q12. Can the sensitivity of the kit assay be increased?

For samples with low concentrations of analyte the sample volume used in the kit assay can be increased to increase sensitivity. When doing this the water volume is adjusted to retain the same final assay volume. This is critical for the manual assay format because the assay volume and sample volume are used in the calculation of results.

Q13. When using this kit for quantitative analysis what level of accuracy and repeatability can be expected?

The test kit is extremely accurate – at Megazyme the quality control criteria for accuracy and repeatability is to be within 2% of the expected value using pure analytes.

However, the level of accuracy is obviously analyst and sample dependent.

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a-D-葡萄糖醛酸酶检测试剂盒 α-Glucuronidase Assay Kit 货号:K-AGLUA Megazyme中文站

a-D-葡萄糖醛酸酶检测试剂盒

英文名:α-Glucuronidase Assay Kit

货号:K-AGLUA

规格:50 Assays per kit

The Alpha-D-Glucuronidase test kit is a simple, reliable and accurate method for the measurement and analysis of alpha-D-glucuronidase in various enzyme preparations. Containing and aldouronic acid substrate and an α-D-glucuronidase control.
Suitable for manual and microplate formats.

UV-method for the measurement of α-D-Glucuronidase in
various enzyme preparations

Principle:
(α-D-glucuronidase)
(1) Aldouronic acid (tri:tetra:penta) + H2O →
β-(1,4)-D-xylo-oligosaccharides + D-glucuronic acid

(uronate dehydrogenase; UDH)
(2) D-Glucuronic acid + NAD+ + H2O → D-glucarate + NADH + H+

Kit size: 50 assays (manual) / 200 (microplate)
Method: Spectrophotometric at 340 nm
Reaction time: ~ 25 min
Detection limit: 17 mU/mL
Application examples:
Enzyme preparations and other materials
Method recognition: Novel method

Advantages

  • Very competitive price (cost per test)
  • All reagents stable for > 2 years as supplied
  • Simple format
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
  • Standard included
  • Suitable for manual, microplate and auto-analyser formats

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木聚糖酶检测试剂盒 Xylanase Assay Kit (Azo-Wax) 货号:K-AZOWAX Megazyme中文站

木聚糖酶检测试剂盒

英文名:Xylanase Assay Kit (Azo-Wax)

货号:K-AZOWAX

规格:200 assays per kit

产品名称:木聚糖酶检测试剂盒
英文名称:Xylanase Assay Kit (Azo-Wax)

型号规格:200次
品牌:爱尔兰Megaeyme公司

自然界中,D-木糖以多糖形式如木聚糖,阿拉伯糖基木聚糖,GAX(glucuronoarabinoxylan),木葡聚糖和XGA(xylogalacturonan)存在。一些海藻中也存在交联D-木糖,据认定一些相近多糖构成洋车前子胶的骨架。番石榴,梨,黑莓,罗甘莓,悬钩子,芦荟维拉胶,褐藻,紫锥菊,乳香树,花椰菜,菠菜,茄子,豌豆,青豆,秋葵,卷心菜和谷物中存在游离的D-木糖。D-木糖用于小肠疾病诊断中的吸收实验,如食品中营养,维生素和矿物质吸收障碍。D-木糖可以被小肠正常轻松地吸收,当患有这些疾病时,D-木糖不能被吸收,血液和尿中的含量很低。D-木糖检测可以帮助确定儿童无法增重的原因,尤其是那些进食足够的儿童。如果在多糖中, D-木糖与其他糖类的比例已知,就可以通过酸水解测定D-木糖的浓度进而定量多糖。木聚糖是一些多糖的主要部分,这些多糖水解后的可发酵糖类可用于生产生物燃料。

原理

D-木糖的α-和β-变旋异构形式互换由木糖变旋酶(XMR)催化(1)。

在pH7.5下,β-D-木糖由β-木糖脱氢酶(β-XDH)催化,被NAD+氧化为D-木糖酸。

The Xylanase (Azo-Wax) test kit is suitable for the measurement and analysis of endo-1,4-β-D-xylanase in enzyme preparations, bread improver mixtures and animal feeds. Containing Azo-wheat arabinoxylan and a Trichoderma sp. xylanase control.

Colourimetric method for the determination of Xylanase in feed,
foodstuffs and other materials

Principle:
(β-xylanase)
(1) Azo-WAX + H2O → Azo-WAX fragments
(insoluble in aqueous alcohol) (soluble in aqueous alcohol)

Kit size: 200 assays
Method: Based on use of Azo-WAX reagent (590 nm)
Total assay time: ~ 45 min
Detection limit: 0.2 U/mL of assay solution
Application examples:
Animal feeds, enzyme preparations, bread improver mixtures and
other materials
Method recognition:
Used widely in the feed industry

Advantages

  • Very cost effective
  • All reagents stable for > 2 years
  • Only test kit available
  • Simple format
  • Standard included

 

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用于测定谷类植物和麦芽中的β-淀粉酵素的试剂盒(RACI标准法) β-amylase Assay (Betamyl-3) 货号:K-BETA3 Megazyme中文站

用于测定谷类植物和麦芽中的β-淀粉酵素的试剂盒(RACI标准法)

英文名:β-amylase Assay (Betamyl-3)

货号:K-BETA3

规格:100/200 assays per kit

For the specific measurement of β-amylase in malt flour. Content: 100/200 assays per kit
 

Colourimetric method for the determination of β-Amylase in
cereal grains, malt, food, beverages and fermentation products

Principle:
(β-amylase)
(1) G3-β-PNP + H2O → G2 + G-β-PNP

(β-glucosidase)
(2) G-β-PNP + H2O → D-glucose + PNP

(alkaline solution)
(3) p-Nitrophenol → phenolate ion (yellow colour)
Note: PNP = 4-nitrophenol

Kit size: 100 / 200 assays
Method: Spectrophotometric at 400 nm
Reaction time: ~ 10 min
Detection limit: 0.05 U/mL of sample solution
Application examples:
Cereal flours, malts and other materials
Method recognition:
Modification of RACI (Standard Method)

Advantages

  • Very cost effective
  • All reagents stable for > 2 years as supplied
  • Only enzymatic kit available
  • Very specific
  • Simple format
  • Rapid reaction
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
  • Standard included

 Q1. Should the pH of the sample be adjusted even for samples in acidic media?

The pH of the assay solution after the sample is added should be the same as that of the assay buffer that is supplied with the kit.
Low sample volumes (e.g. 0.1 mL) are not likely to affect the pH of the assay solution and therefore may not require pH adjustment.
Samples above 0.1 mL are more likely to affect the pH of the assay solution and therefore the pH of these samples should be adjusted as described in the data booklet, prior to addition to the assay.

Q2. There is an issue with the performance of the kit; the results are not as expected.

If you suspect that the Megazyme test kit is not performing as expected such that expected results are not obtained please do the following:

  1. Ensure that you have tested the standard sample that is supplied with the Megazyme test kit.
  2. Send the results of the kit standard, blank samples and the results obtained for your sample,  in the relevant MegaCalc spreadsheet (if available) to Megazyme (cs@megazyme.com). Where available the relevant MegaCalc spreadsheet can be downloaded from where the product appears on the Megazyme website.
  3. State the kit lot number being used (this is found on the outside of the kit box).
  4. State which assay format was used (refer to the relevant page in the kit booklet if necessary).
  5. State exact details of any modifications to the standard procedure that is provided by Megazyme.
  6. State the sample type and describe the sample preparation steps if applicable.

内切纤维素酶检测试剂盒[CELLG3方法] Cellulase Assay Kit (CELLG3 Method) 货号:K-CELLG3 Megazyme中文站

内切纤维素酶检测试剂盒[CELLG3方法]

英文名:Cellulase Assay Kit (CELLG3 Method)

货号:K-CELLG3

规格:180 / 360 assays per kit / 720 (auto-analyser)

 纤维素酶是一种重要的酶产品,是一种复合酶,主要由外切β-葡聚糖酶、内切β-葡聚糖酶和β-葡萄糖苷酶等组成,还有很高活力的木聚糖酶。

提供内切纤维素酶检测试剂盒,色度法(400 nm)测定酶制品和发酵产品中的纤维素酶(内切-1,4-β-葡聚糖酶)。

内切纤维素酶检测试剂盒(CELLG3方法) K-CELLG3

使用高纯度β-葡萄糖苷酶和苯亚甲基阻断,2 – 氯-4 – 硝基苯基-β-Dcellotrioside(BClPNPβ-G3)。 β-葡萄糖苷酶的能力确保测定的可靠的最大的灵敏度。BClPNPβ-G3的水解为苯亚甲基,通过纤维素酶阻断纤维二糖和2-氯-4 -硝基苯基-β-D-葡萄糖, 2-Cl-4-硝基苯基-β-D-葡萄糖通过β-葡萄糖苷酶立即裂解为D-葡萄糖和游离的2 – 氯-4 – 硝基苯酚(ClPNP)。

反应时间:16min

适用样品:酶制品和发酵产品

优点:

价格低廉(每次检测成本)

特异性高

方法简单

包含标准品

 

 The CELLG3 assay reagent for the measurement of endo-cellulase (endo-1,4-β-glucanase) contains two components; 

1) 4,6-O-benzylidene-2-chloro-4-nitrophenyl-β-D-cellotrioside (BCNPG3) and 2) thermostable β-glucosidase. The benzylidene blocking group prevents any hydrolytic action by the β-glucosidase on BCNPG3.  Incubation with an endo-cellulase generates a non-blocked colourimetric oligosaccharide that is rapidly hydrolysed by the ancillary β-glucosidase.  The rate of formation of 2-chloro-4-nitrophenol is therefore directly related to the hydrolysis of BCNPG3 by the endo-cellulase.  The reaction is terminated and the phenolate colour is developed on addition of Tris buffer solution (pH 9.0).
Please note that a new assay kit (K-CELLG5) is now available for the measurement of endo-cellulase.  The CELLG5 reagent contains a cellopentaose core and exhibits vastly improved sensitivity for some cellulases.  In addition, the exchange of the benzylidene blocking group in CELLG3 for 3-keto-butylidene in CELLG5 improves the substrate’s water solubility significantly, allowing for a reduction in the concentration of DMSO required in the assay.  As DMSO is known to inhibit certain cellulases, this is another benefit in using CELLG5.  Megazyme now recommends the use of K-CELLG5 for all assays for the measurement of endo-cellulase.

Colourimetric method for the determination of 
endo
-1,4-β-glucanase (cellulase) in enzyme preparations and fermentationproducts

Principle:
                                       (endo-1,4-β-glucanase)
(1) 3-Ketobutylidene-G5-β-PNP + H2O → Blocked-GX + G(5-X)-β-PNP

                (thermostable β-glucosidase)
(2) G(5-X)-β-PNP + H2O → D-glucose + PNP

      (alkaline solution)
(3) PNP → phenolate ion (yellow colour)
Note: PNP = 4-nitrophenol

Kit size:
K-CELLG5-4V 120 / 240 assays (manual) / 480 (auto-analyser)
                                        or
K-CELLG5-2V 60 / 120 assays (manual) / 240 (auto-analyser)

Method:                         Spectrophotometric at 400 nm
Total assay time:           10 min
Detection limit:                3.5 x 10-4 U/mL
Application examples:
Fermentation broths, industrial enzyme preparations, biofuels research
Method recognition:     Novel method


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内切纤维素酶检测试剂盒[CELLG5方法] Cellulase Assay Kit (CELLG5 Method) 货号:K-CELLG5-2V Megazyme中文站

内切纤维素酶检测试剂盒[CELLG5方法]

英文名:Cellulase Assay Kit (CELLG5 Method)

货号:K-CELLG5-2V

规格:60 / 120 assays (manual) / 240 (auto-analyser)

The CELLG5 assay reagent for the measurement of
endo-cellulase (endo-1,4-β-glucanase) contains two components;
1) 4,6-O-(3-Ketobutylidene)-4-nitrophenyl-β-D-cellopentaoside (BPNPG5) and 2) thermostable β-glucosidase. The ketone blocking group prevents any hydrolytic action by the
β-glucosidase on BPNPG5. Incubation with an endo-cellulase generates a non-blocked colourimetric oligosaccharide that is rapidly hydrolysed by the ancillary β-glucosidase. The rate of formation of 4-nitrophenol is therefore directly related to the hydrolysis of BPNPG5 by the endo-cellulase. The reaction is terminated and the phenolate colour is developed on addition of Tris buffer solution (pH 9.0).

 

Colourimetric method for the determination of
endo
-1,4-β-glucanase (cellulase) in enzyme preparations and fermentation products

Principle:
(endo-1,4-β-glucanase)
(1) 3-Ketobutylidene-G5-β-PNP + H2O → Blocked-GX + G(5-X)-β-PNP

(thermostable β-glucosidase)
(2) G(5-X)-β-PNP + H2O → D-glucose + PNP

(alkaline solution)
(3) PNP → phenolate ion (yellow colour)
Note: PNP = 4-nitrophenol

Kit size:
K-CELLG5-4V 120 / 240 assays (manual) / 480 (auto-analyser)
or
K-CELLG5-2V 60 / 120 assays (manual) / 240 (auto-analyser)

Method: Spectrophotometric at 400 nm
Total assay time: 10 min
Detection limit: 3.5 x 10-4 U/mL
Application examples:
Fermentation broths, industrial enzyme preparations, biofuels research
Method recognition: Novel method

Advantages

  • Very cost effective
  • All reagents stable for > 4 years
  • Completely specific for cellulase (endo-1,4-glucanase)
  • Generally applicable and highly sensitive
  • Simple format. Well suited to automation
  • Standard included

 

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内切纤维素酶检测试剂盒[CELLG5方法] Cellulase Assay Kit (CELLG5 Method) 货号:K-CELLG5-4V Megazyme中文站

内切纤维素酶检测试剂盒[CELLG5方法]

英文名:Cellulase Assay Kit (CELLG5 Method)

货号:K-CELLG5-4V

规格:120 / 240 assays (manual) / 480 (auto-analyser)

The CELLG5 assay reagent for the measurement of
endo-cellulase (endo-1,4-β-glucanase) contains two components;
1) 4,6-O-(3-Ketobutylidene)-4-nitrophenyl-β-D-cellopentaoside (BPNPG5) and 2) thermostable β-glucosidase. The ketone blocking group prevents any hydrolytic action by the
β-glucosidase on BPNPG5. Incubation with an endo-cellulase generates a non-blocked colourimetric oligosaccharide that is rapidly hydrolysed by the ancillary β-glucosidase. The rate of formation of 4-nitrophenol is therefore directly related to the hydrolysis of BPNPG5 by the endo-cellulase. The reaction is terminated and the phenolate colour is developed on addition of Tris buffer solution (pH 9.0).

 

Colourimetric method for the determination of
endo
-1,4-β-glucanase (cellulase) in enzyme preparations and fermentation products

Principle:
(endo-1,4-β-glucanase)
(1) 3-Ketobutylidene-G5-β-PNP + H2O → Blocked-GX + G(5-X)-β-PNP

(thermostable β-glucosidase)
(2) G(5-X)-β-PNP + H2O → D-glucose + PNP

(alkaline solution)
(3) PNP → phenolate ion (yellow colour)
Note: PNP = 4-nitrophenol

Kit size:
K-CELLG5-4V 120 / 240 assays (manual) / 480 (auto-analyser)
or
K-CELLG5-2V 60 / 120 assays (manual) / 240 (auto-analyser)

Method: Spectrophotometric at 400 nm
Total assay time: 10 min
Detection limit: 3.5 x 10-4 U/mL
Application examples:
Fermentation broths, industrial enzyme preparations, biofuels research
Method recognition: Novel method

Advantages

  • Very cost effective
  • All reagents stable for > 4 years
  • Completely specific for cellulase (endo-1,4-glucanase)
  • Generally applicable and highly sensitive
  • Simple format. Well suited to automation
  • Standard included

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Alpha淀粉酶/液化酶/淀粉糖化酶检测试剂盒 Ceralpha: Alpha-Amylase Assay Kit 货号:K-CERA Megazyme中文站

Alpha淀粉酶/液化酶/淀粉糖化酶检测试剂盒

英文名:Ceralpha: Alpha-Amylase Assay Kit

货号:K-CERA

规格:100 assays per kit

AOAC Method 2002.01, AACC Method 22.02.01, ICC Standard No. 303, RACI Standard Method, CCFRA Flour Testing Working Group Method 0018. For the specific measurement of α-amylase in cereal grains and fermentation broths (fungal and bacterial).

Alpha淀粉酶/液化酶/淀粉糖化酶检测试剂盒

Q1. We would like to obtain an assay capable of quantifying trace amounts of alpha-amylase activity in the pullulanase preparation. Please let me know if Megazyme has any suitable assay available.

We think the Ceralpha Reagent would probably be best for this application.

Q2. I am interested in the Amylazyme and Ceralpha available from Megazyme. Are they applicable not only for cereal and microbial alpha-amylase but also porcine pancreatic alpha-amylase?

The Ceralpha method is excellent for all alpha-amylases and will work fine for porcine pancreas alpha-amylase at pH 6.9.

Q3. We would like to know where the coefficient of 18.1 (EmM of p-nitrophenol) comes from? What does it mean?

This is the extinction coefficient, i.e. absorbance of a 1 mM solution of p-nitrophenol in a 1 cm light path at 400 nm.

Q4. I wish to measure the enzyme activity of alpha-amylase in bread after cooking. Will your Ceralpha Amylase Kit work on cooked breads? If so, do I need to alter the procedure?

The level of alpha-amylase in wheat flour is quite low, so in bread baked from this flour it is likely to be much lower.  The Ceralpha test is very sensitive, but you can increase sensitivity by increasing incubation time up to several hours (6 hours).  This should allow measurement of the enzyme if there is any there.

Q5. Any recommendations on using your Ceralpha alpha-amylase test at a low pH (pH 3-4)?

Cereal alpha-amylase assay reagent cannot be used at pH 3-4.  However, you can perform a similar assay using blocked p-nitophenol maltoheptaoside (BPNPG7). Basically dissolve the BPNPG7 vial contents in 10 mL with water.  Incubate 0.2 mL of your enzyme (buffered) with 0.2 mL of BPNPG7 for 10 min at a set temperature. Terminate the reaction by heating in a boiling water bath for 2 min.  Then add 0.2 mL of alpha-glucosidase (E-TSAGL; at 10 U/mL in 0.2 M tris buffer, pH 7.0) and incubate for 10 min.  Add 3 mL stopping reagent (Trizma base pH 8.5) and read colour at 400 nm.

Q6. Can the Ceralpha kit distinguish between different forms of alpha-amylase?

The Ceralpha Kit does not distinguish between the different forms of alpha amylase, however some idea of relative levels of fungal and bacterial alpha-amylase can be obtained by performing the assay at different pH values.  Please refer to the Ceralpha kit booklet on this web site.

Q7. In order to use the Ceralpha kit to test for the presence of alpha-amylase, does one have to mill the wheat and if so can I grind the grain in a blender?

Cereal grains do need to be milled to allow effective extraction of alpha-amylase (and other constituents).  We recommend milling to pass a 0.5 mm screen.  You may have some success with a cheap coffee grinder.  However, the reproducibility will not be as good (it may be adequate for your requirements).

Q8. Can the Ceralpha reagent be used at high temperatures?

The Ceralpha method is routinely run at 40˚C, but it can be used at temperatures up to 60˚C.  If it is run at higher temperatures (up to 60˚C) then higher activity values will be obtained.  This assumes that the enzymes being analysed (alpha-amylase) are stable at these higher temperatures.  If you need to measure activity at temperatures above 60˚C you can use the Megazyme Amylazyme tablets or Red Starch.

Q9. Can Ceralpha Method determine the alpha-amylase activity in a protease preparation?

The standard method should be fine.  You can best check this by performing a time course hydrolysis of the alpha-amylase.  This should be linear if the alpha-glucosidase in the kit reagent is not attacked by the protease.

Q10. Can the internal glycosidic linkages only be cleaved by alpha-amylase?

Yes, only alpha-amylase can cleave the internal glycosidic bonds.

Q11. After alpha-amylase cleaves the internal glycosidic linkage, can beta-amylase or glucoamylase accelerate the hydrolysis reaction of the released p-nitrophenyl maltosaccharide?

When alpha-amylase cleaves the glycosidic linkage in the blocked substrate, any other enzymes in the extract do not accelerate the hydrolysis to give free p-nitrophenol.  The reason for this is that the level of thermostable alpha-glucosidase in the substrate mixture is saturating.

Q12. Can Rice alpha-amylase be analysed by Ceralpha kit?

Rice alpha-amylase can be assayed with Ceralpha kit.  Alpha-glucosidase in the rice extract will not interfere.

Q13. Why is extraction conducted at room temperature or 40˚C, and not at 4˚C?

We have found that we get effective extraction from a range of cereal flours at room temperature (approx. 22˚C) and that the enzyme was very stable at this temperature for several hours.  For wheat flour, the optimal temperature for extraction is 40˚C (refer to the Ceralpha Booklet).

Q14. Can the Ceralpha method be used to determine alpha-amylase from oat flour extracts?

The Ceralpha method can be used for oats.  The only likely problem is the high viscosity of the extract.  If the extract looks very viscous, double the ratio of extraction buffer to flour.  Allow for this in the calculations.  If the absorbances are low then increase the incubation time to say 30 minutes.  Then allow for this increased time of incubation in the calculations.

Q15. Is there a method to increase the sensitivity of assay regarding the measurement of alpha-amylase in fermentation broth?

We suggest that the fermentation broth is simply adjusted to 5.0 with acid or base and this can be used directly in the assay. The sensitivity can then be increased by increasing incubation time up to 4 hours.  Appropriate adjustments then need to be made to the calculations.

Q16. Can the reaction blank be used to zero the spectrophotometer directly?

You can zero the spectrophotometer directly with the blank.  However, it is wise to know how high the blank is to ensure that the substrate is OK, i.e. the blank is usually 0.05-0.06.  If it is above 0.1, it has probably been contaminated with some alpha-amylase enzyme.

Q17. When the absorbance is between 1.0–1.5, can the solution be diluted directly with an additional 3.0 mL of distilled water to cut the absorbance in half before reading it?

If the absorbance value is above 1.2 then the assay will be limited by the amount of available substrate.  Thus, you cannot simply dilute the colour.

Q18. I have immobilised alpha-amylase onto glass bead and now I want to test the activity, can the Ceralpha kit test for this?

To measure alpha-amylase on glass beads I would recommend the use of the Ceralpha method.  The kit is complete and extremely easy to use.  Perhaps to assay, you may freeze dry a small sample of the beads and then weigh an appropriate amount (say 5 mg) into a test tube.  Add 0.2 mL of buffer pH 5-6 and 0.2 mL Ceralpha reagent.  Incubate at 40˚C for 10 min.

Q19. Is it possible to measure acid-stable alpha-amylase by the Ceralpha Kit?

The Ceralpha method should be fine for acid stable alpha-amylase.  However, assays should be performed in the pH range of 5.2 to 7.5.

Q20. Can you tell me the application of the Amylazyme Tabs and Ceralpha Kit to residual enzymes in finished bakery products, particularly in sweet goods?

For measurement of residual enzymes in bakery products, you should use the Amylazyme method rather than the Ceralpha method.  With the standard assay formats available, the sensitivity of the Amylazyme method is 10-20 times greater than can be achieved with the Ceralpha method.  You will need this greater sensitivity to measure the extremely low levels of activity present.

Q21. What would be an expected level of Ceralpha units in germinated or malted barley?

The level of alpha-amylase in malted barley is about 150 Ceralpha units per gram, which would be the expected level in germinated barley grain.

Q22. Should the pH of the sample be adjusted even for samples in acidic media?

The pH of the assay solution after the sample is added should be the same as that of the assay buffer that is supplied with the kit.
Low sample volumes (e.g. 0.1 mL) are not likely to affect the pH of the assay solution and therefore may not require pH adjustment.

葡萄糖氧化酶检测试剂盒 Glucose Oxidase Assay Kit 货号:K-GLOX Megazyme中文站

葡萄糖氧化酶检测试剂盒

英文名:Glucose Oxidase Assay Kit

货号:K-GLOX

规格:200 assays (manual) / 2000 assays (microplate)

The Glucose Oxidase assay kit is a simple procedure for the rapid and reliable measurement and analysis of glucose oxidase activity in industrial enzyme preparations and bread improver mixtures.

Glucose Oxidase assay kit is suitable for manual, auto-analyser and microplate formats.
 

Colourimetric method for the determination of Glucose Oxidase
in foodstuffs and fermentation products

Principle:
(glucose oxidase)
(1) D-Glucose + H2O + O2 → D-glucono-δ-lactone + H2O2

(peroxidase)
(2) 2H2O2 + p-hydroxybenzoic acid + 4-aminoantipyrine →
quinoneimine + 4H2O

Kit size: 200 assays (manual) / 2000 (microplate)
/ 1960 (auto-analyser)
Method: Spectrophotometric at 510 nm
Reaction time: ~ 20 min
Detection limit: 10 U/L
Application examples:
Enzyme preparations, and other materials (e.g. biological cultures,
samples, etc.)
Method recognition: Novel method

Advantages

  • Very competitive price (cost per test)
  • All reagents stable for > 12 months after preparation
  • Simple format
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
  • Standard included
  • Suitable for manual, microplate and auto-analyser formats

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麦芽淀粉酶检测试剂盒 Malt Amylase Assay Kit 货号:K-MALTA Megazyme中文站

麦芽淀粉酶检测试剂盒

英文名:Malt Amylase Assay Kit

货号:K-MALTA

规格:100 assays (50 of each) per kit

The Malt Amylase test kit is suitable for the specific measurement and analysis of α-amylase and of β-amylase in malt flour.

Colourimetric method for the determination of α-Amylase and
β-Amylase in cereal grains, malt, food, beverages and
fermentation products

Principle:
(1) α-Amylase is measured using the “Ceralpha” Method as used
in K-CERA

(2) β-Amylase is measured using the “Betamyl-3” Method as used
in K-BETA3

Kit size: 50 assays of each
Method: Spectrophotometric at 400 nm
Reaction time: ~ 20 min (Ceralpha Method)
~ 10 min (Betamyl-3 Method)
Detection limit: 0.05 U/mL
Application examples:
Cereal flours, malts, fermentation broths and other materials
Method recognition:
“Ceralpha” Method: AOAC (Method 2002.01), AACC (Method
22-02.01), ICC (Standard No. 303), RACI
(Standard Method) and CCFRA (Flour Testing
Working Group Method 0018)
“Betamyl-3” Method: RACI (Standard Method)

Advantages

  • Very cost effective
  • All reagents stable for > 2 years as supplied
  • Only enzymatic kit available (Beta-Amylase)
  • Very specific
  • Simple format
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
  • Standard included

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Beta葡聚糖酶[麦芽和微生物]检测试剂盒 β-Glucanase Assay Kit (Malt and Microbial) 货号:K-MBGL Megazyme中文站

Beta葡聚糖酶[麦芽和微生物]检测试剂盒

英文名:β-Glucanase Assay Kit (Malt and Microbial)

货号:K-MBGL

规格:100 assays per kit

β-Glucanase (Malt and Microbial) Assay Kit is suitable for the measurement and analysis of malt and bacterial β-glucanase and endo-1,4-β-glucanase.

Colourimetric method for the determination of β-Glucanase
in malt, foodstuffs and fermentation products

Principle:
(β-glucanase)
(1) Azo-Barley β-glucan (polymer) → Azo-barley β-glucan
(fragments)

(2) Add alcohol; centrifuge to remove polymeric Azo-barley
β-glucan

(3) Measure the absorbance of the supernatant solution

Kit size: 100 assays
Method: Spectrophotometric at 590 nm
Reaction time: ~ 30 min
Detection limit: 100 U/kg of malt
Application examples:
Malt extracts, wort, beer and other materials
Method recognition:
RACI (Standard Method)

Advantages

  • Very cost effective
  • All reagents stable for > 2 years during use
  • Only kit available
  • Very specific
  • Simple format
  • Standard included

Q1. We have measured the molecular weight of beta-glucan originating from barley ordered from your company. Can you please tell us which method you have used for measuring molecular weight?

The MW’s were determined by Multiangle laser light scattering technique.

Q2. I have purchased barley beta-glucan (lot 30108) and carob galactomannan low viscosity (lot 30702) and would like to know what else there might be in these substrates.

Barley beta-glucan lot 30108 would contain about 3-4% arabinoxylan (this was produced in 1993).  Material supplied post 1995 contains < 0.5.% arabinoxylan. Carob galactomannan is quite pure (> 96%).

Q3. Although not specified on the beta-glucan data sheet, is the ratio of 1-3 to 1-4 bonds measured? If so, what is the ratio and would you expect it to remain standardised over a number of different batches?

The content of 1,3 bonds in barley beta-glucan is about 32% (from literature).  We would not expect this to change much (if at all) over different batches.

Q4. Both substrates, barley beta-glucan and wheat arabinoxylan, are standardised to a specific viscosity, e.g. 23-24 cSt. Are the substrates adjusted to give this viscosity?

The substrates are enzymically treated to yield the required viscosity (20 ~ 30 cSt). Modification of the viscosity of beta-glucan is required for IOB viscometric malt beta-glucanase test (which works well).

Q5. Should the pH of the sample be adjusted even for samples in acidic media?

The pH of the assay solution after the sample is added should be the same as that of the assay buffer that is supplied with the kit.
Low sample volumes (e.g. 0.1 mL) are not likely to affect the pH of the assay solution and therefore may not require pH adjustment.
Samples above 0.1 mL are more likely to affect the pH of the assay solution and therefore the pH of these samples should be adjusted as described in the data booklet, prior to addition to the assay.

Q6. There is an issue with the performance of the kit; the results are not as expected.

If you suspect that the Megazyme test kit is not performing as expected such that expected results are not obtained please do the following:

  1. Ensure that you have tested the standard sample that is supplied with the Megazyme test kit.
  2. Send the results of the kit standard, blank samples and the results obtained for your sample, in the relevant MegaCalc spreadsheet (if available) to Megazyme (cs@megazyme.com). Where available the relevant MegaCalc spreadsheet can be downloaded from where the product appears on the Megazyme website.
  3. State the kit lot number being used (this is found on the outside of the kit box).
  4. State which assay format was used (refer to the relevant page in the kit booklet if necessary).
  5. State exact details of any modifications to the standard procedure that is provided by Megazyme.
  6. State the sample type and describe the sample preparation steps if applicable.

Q7. We want to set up reducing-sugar assays for beta-glucanase and xylanase from different fungal origins. There are three kinds in your catalogue – high viscosity, medium viscosity and low viscosity. Which do you recommend for the glucanase assay.

For glucanase assay we recommend the medium viscosity beta-glucan.  We now offer low, medium and high viscosity wheat arabinoxylans, and we think that the low viscosity material will be best (easiest) to use in the reducing-sugar assay. 

普鲁兰酶/极限醍醐精检测试剂盒 Pullulanase/Limit-Dextrinase Assay Kit (PULLG6 Method) 货号:K-PULLG6 Megazyme中文站

普鲁兰酶/极限醍醐精检测试剂盒

英文名:Pullulanase/Limit-Dextrinase Assay Kit (PULLG6 Method)

货号:K-PULLG6

规格:100 assays (manual)

PULLG6 assay for the measurement of pullulanase employs a water soluble defined substrate, namely 4,6-O-benzylidene-4-nitrophenyl-63-α-D-maltotriosyl-maltotriose (BPNPG3G3), coupled with the ancillary enzymes α-glucosidase and β-glucosidase. Upon hydrolysis of the substrate at the 1,6-α-linkage by pullulanase or limit-dextrinase, the released 4-nitrophenyl-β-maltotrioside is immediately hydrolysed to glucose and 4-nitrophenol by the concerted action of the α-glucosidase and β-glucosidase enzymes in the reagent mixture. The reaction is terminated and phenolate ions are developed by addition of dilute alkali. The absorbance is read at 400 nm and the value obtained correlates directly with pullulanase activity.

Colourimetric method for the determination of pullanase
or limit-dextrinase

Principle:
(pullulanase/limit-dextrinase)
(1) Benzylidene-G3-(α-1,6)-G3-β-PNP + H2O → Benzylidene-G3
+ G3-β-PNP

(thermostable α-glucosidase and β-glucosidase)
(2) G3-β-PNP + H2O → D-glucose + PNP

(alkaline solution)
(3) PNP → phenolate ion (yellow colour)

Note: PNP = 4-nitrophenol

Kit size: 100 assays
Method: Spectrophotometric at 400 nm
Total assay time: 10 min for pullanase preparations
30 min for malt extracts containing
limit-dextrinase
Detection limit: 0.18 U/mL for pullulanase preparations
(50-fold dilution)
0.01 U/g for limit dextrinase in milled malt
Application examples: Assay of microbial pullulanase preparations
Measurement of limit-dextrinase in
malt extracts
Method recognition: Novel method

Advantages

  • High sensitivity
  • Suitable for manual and auto-analyser formats
  • No transglycosylation interference
  • Very cost effective
  • All reagents stable for > 1 year after preparation
  • Very specific

  • Simple format

  • Standard included

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木聚糖酶检测试剂盒 Xylanase Assay Kit (Xylazyme AX) 货号:K-XYLS Megazyme中文站

木聚糖酶检测试剂盒

英文名:Xylanase Assay Kit (Xylazyme AX)

货号:K-XYLS

规格:200 assays per kit

The Xylanase (Xylazyme AX) test kit is used for the measurement and analysis of endo-1,4-β-D-xylanase in enzyme preparations, bread improver mixtures and animal feeds. Contains Xylazyme AX Tablets and xylanase enzyme controls (A. niger and Trichoderma longibrachiatum).

Colourimetric method for the determination of Xylanase in feed,
foodstuffs and other materials

Principle:
(β-xylanase)
(1) Xylazyme AX (water insoluble) + H2O → water soluble dyed xylan fragments

Kit size: 200 assays
Method: Based on use of Xylazyme AX tablets (590 nm)
Total assay time: ~ 45 min
Detection limit: 0.02 U/mL of assay solution
Application examples:
Animal feeds, enzyme preparations, bread improver mixtures and
other materials
Method recognition:
Used widely in the feed industry

Advantages

  • Very cost effective
  • All reagents stable for > 2 years during use
  • Only test kit available
  • Simple format
  • Standard included

 

 Q1. Should the pH of the sample be adjusted even for samples in acidic media?

The pH of the assay solution after the sample is added should be the same as that of the assay buffer that is supplied with the kit.
Low sample volumes (e.g. 0.1 mL) are not likely to affect the pH of the assay solution and therefore may not require pH adjustment.
Samples above 0.1 mL are more likely to affect the pH of the assay solution and therefore the pH of these samples should be adjusted as described in the data booklet, prior to addition to the assay.

Q2. There is an issue with the performance of the kit; the results are not as expected.

If you suspect that the Megazyme test kit is not performing as expected such that expected results are not obtained please do the following:

  1. Ensure that you have tested the standard sample that is supplied with the Megazyme test kit.
  2. Send the results of the kit standard, blank samples and the results obtained for your sample, in the relevant MegaCalc spreadsheet (if available) to Megazyme (cs@megazyme.com). Where available the relevant MegaCalc spreadsheet can be downloaded from where the product appears on the Megazyme website.
  3. State the kit lot number being used (this is found on the outside of the kit box).
  4. State which assay format was used (refer to the relevant page in the kit booklet if necessary).
  5. State exact details of any modifications to the standard procedure that is provided by Megazyme.
  6. State the sample type and describe the sample preparation steps if applicable.

α淀粉酶SD检测试剂盒(高感光度法) α-Amylase SD Assay Kit (High Sensitivity Method) 货号:K-AMYLSD Megazyme中文站

α淀粉酶SD检测试剂盒(高感光度法)

英文名:α-Amylase SD Assay Kit (High Sensitivity Method)

货号:K-AMYLSD

规格:160 / 320 assays (manual) / 640 assays (auto-analyser)

The Amylase SD Method: A highly sensitive colourimetric method for the determination of α-amylase in sprout damaged wheat grain (also known as pre-harvest sprouting or weather damaged wheat grain) and “late maturity α-amylase” wheat grain.

Advantages

  • Extremely high sensitivity – 2.4-fold increase over Ceralpha (K-CERA)
  • Very cost effective
  • All reagents stable for > 2 years after preparation
  • Very specific
  • Simple format
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
  • Standard included

  • Suitable for Maual and auto-analyser formats

Highly sensitive colourimetric method for the determination of α-Amylase in sprout damaged grain

Principle:
(α-amylase)
(1) Ethylidene-G7-α-PNP + H2O → Ethylidene-GX + G(7-X)-α-PNP

(thermostable α-glucosidase)
(2) G(7-X)-α-PNP + H2O → D-glucose + PNP

(alkaline solution)
(3) PNP → phenolate ion (yellow colour)
Note: PNP = 4-nitrophenol

Kit size: 160 / 320 assays (manual) / 640 (auto-analyser)
Method: Spectrophotometric at 400 nm
Total assay time: ~ 5 min
Detection limit: 0.05 U/mL
Application examples: Sprout damaged wheat grain
Method recognition: Novel method

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Beta葡聚糖[酵母和蘑菇]检测试剂盒 β-Glucan Assay Kit (Yeast & Mushroom) 货号:K-YBGL Megazyme中文站

Beta葡聚糖[酵母和蘑菇]检测试剂盒

英文名: β-Glucan Assay Kit (Yeast & Mushroom)

货号:K-YBGL

规格:100 assays per kit

For the measurement of 1,3:1,6-ß-glucan and α-glucan in yeast preparations. Content: 100 assays per kit

Colourimetric method for the determination of Yeast and 
Mushroom β-Glucan in yeast, mushroom, foodstuffs and 
other materials

Principle:
                                                              (conc. HCl, 30°C, 45 min)
(1) 1,3:1,6-β-Glucan + 1,3-β-glucan + α-glucan + H2O → 
                                                                        soluble glucan

                         (1.3 M HCl, 100°C, 2 h)
(2) Soluble glucan + H2O → D-glucose + laminarisaccharides (trace)

                  (exo-1,3-β-glucanase + β-glucosidase)
(3) Laminarisaccharides + H2O → D-glucose

                              (glucose oxidase)
(4) D-Glucose + H2O + O2 → D-gluconate + H2O2

                                                                                (peroxidase)
(5) 2H2O2 + p-hydroxybenzoic acid + 4-aminoantipyrine → 
                                                                        quinoneimine + 4H2O

                       (amyloglucosidase)
(6) α-Glucan + H2O    →    D-glucose

Kit size:                            100 assays
Method:                            Spectrophotometric at 510 nm
Total assay time:              ~ 100 min
Detection limit:                 1-100% of sample weight
Application examples:
Yeast preparations, mushroom preparations and other materials
Method recognition:        Novel method

Advantages

  • Very cost effective
     
  • All reagents stable for > 12 months after preparation
     
  • Only enzymatic kit available
     
  • Simple format
     
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
     
  • Standard included

 

Q1. There is an issue with the performance of the kit; the results are not as expected.

If you suspect that the Megazyme test kit is not performing as expected such that expected results are not obtained please do the following:

  1. Ensure that you have tested the standard sample that is supplied with the Megazyme test kit.
  2. Send the results of the kit standard, blank samples and the results obtained for your sample,  in the relevant MegaCalc spreadsheet (if available) to Megazyme (cs@megazyme.com). Where available the relevant MegaCalc spreadsheet can be downloaded from where the product appears on the Megazyme website.
  3. State the kit lot number being used (this is found on the outside of the kit box).
  4. State which assay format was used (refer to the relevant page in the kit booklet if necessary).
  5. State exact details of any modifications to the standard procedure that is provided by Megazyme.
  6. State the sample type and describe the sample preparation steps if applicable.

 

Q2. Should the pH of the sample be adjusted even for samples in acidic media?

The pH of the assay solution after the sample is added should be the same as that of the assay buffer that is supplied with the kit.
Low sample volumes (e.g. 0.1 mL) are not likely to affect the pH of the assay solution and therefore may not require pH adjustment.
Samples above 0.1 mL are more likely to affect the pH of the assay solution and therefore the pH of these samples should be adjusted as described in the data booklet, prior to addition to the assay.

Beta葡聚糖[混联]检测试剂盒 β-Glucan Assay Kit (Mixed Linkage) 货号:K-BGLU Megazyme中文站

Beta葡聚糖[混联]检测试剂盒

英文名:β-Glucan Assay Kit (Mixed Linkage)

货号:K-BGLU

规格:100 assays per kit

分析物意义: 大麦和燕麦的主要细胞壁多糖 

Megazyme检测试剂盒优点:反应迅速、试剂稳定, 只有酶检测试剂盒可用。 AOAC方法 995.16; AACC 方法 32-23 

AACC Method 32-23.01, AOAC Method 995.16, EBC Methods 3.11.1, 4.16.1 and 8.11.1, ICC Standard No. 166 and RACI standard method for the measurement of 1,3:1,4-ß-D-glucan in cereal grains, milling fractions, wort and beer. Recommended/Standard procedure of the Association of Official Analytical Chemists (AOAC), American Association of Cereal Chemists (AACC), Royal Australian Chemical Institute (RACI), European Brewing Convention (EBC), and International Association for Cereal Science and Technology (ICC). Content:100 assays per kit

Colourimetric method for the determination of β-Glucan in
cereal grains, feed, foodstuffs, beverages and other materials

Principle:
(lichenase)
(1) β-Glucan + H2O → β-gluco-oligosaccharides

(β-glucosidase)
(2) β-Gluco-oligosaccharides + H2O → D-glucose

(glucose oxidase)
(3) D-Glucose + H2O + O2 → D-gluconate + H2O2

(peroxidase)
(4) 2H2O2 + p-hydroxybenzoic acid + 4-aminoantipyrine →
quinoneimine + 4H2O

Kit size: 100 assays
Method: Spectrophotometric at 510 nm
Total assay time: ~ 100 min
Detection limit: 0.5-100% of sample weight
Application examples:
Oats, barley, malt, wort, beer, food and other materials
Method recognition:
AOAC (Method 995.16), AACC (Method 32-23.01), EBC (Methods
3.10.1, 4.16.1 and 8.13.1), ICC (Standard No. 166), RACI (Standard
Method) and CODEX (Type II Method)

Advantages

  • Very cost effective
  • All reagents stable for > 2 years as supplied
  • Only enzymatic kit available
  • Very specific
  • Simple format
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
  • Standard included

Q1. Should the pH of the sample be adjusted even for samples in acidic media?

The pH of the assay solution after the sample is added should be the same as that of the assay buffer that is supplied with the kit.
Low sample volumes (e.g. 0.1 mL) are not likely to affect the pH of the assay solution and therefore may not require pH adjustment.
Samples above 0.1 mL are more likely to affect the pH of the assay solution and therefore the pH of these samples should be adjusted as described in the data booklet, prior to addition to the assay.

Q2. There is an issue with the performance of the kit; the results are not as expected.

If you suspect that the Megazyme test kit is not performing as expected such that expected results are not obtained please do the following:

  1. Ensure that you have tested the standard sample that is supplied with the Megazyme test kit.
  2. Send the results of the kit standard, blank samples and the results obtained for your sample,  in the relevant MegaCalc spreadsheet (if available) to Megazyme (cs@megazyme.com). Where available the relevant MegaCalc spreadsheet can be downloaded from where the product appears on the Megazyme website.
  3. State the kit lot number being used (this is found on the outside of the kit box).
  4. State which assay format was used (refer to the relevant page in the kit booklet if necessary).
  5. State exact details of any modifications to the standard procedure that is provided by Megazyme.
  6. State the sample type and describe the sample preparation steps if applicable.

Q3. We have measured the molecular weight of beta-glucan originating from barley ordered from your company. Can you please tell us which method have you used for measuring molecular weight?

The MW’s were determined by Multiangle laser light scattering technique.

Q4. I have purchased barley beta-glucan (lot 30108) and carob galactomannan low viscosity (lot 30702) and would like to know what else there might be in these substrates.

Barley beta-glucan lot 30108 would contain about 3-4% arabinoxylan (this was produced in 1993).  Material supplied post 1995 contains < 0.5% arabinoxylan.  Carob galactomannan is quite pure (> 96%).

Q5. Although not specified on the beta-glucan data sheet, is the ratio of 1-3 to 1-4 bonds measured? If so, what is the ratio and would you expect it to remain standardised over a number of different batches?

The content of 1,3 bonds in barley beta-glucan is about 32% (from literature).  We would not expect this to change much (if at all) over different batches.

Q6. Both substrates; barley beta-glucan and wheat arabinoxylan, are standardised to a specific viscosity, e.g. 23-24 cSt. Are the substrates adjusted to give this viscosity?

The substrates are enzymically treated to yield the required viscosity (20 ~ 30 cSt). Modification of the viscosity of beta-glucan is required for IOB viscometric malt beta-glucanase test (which works well).

Q7. We are setting up reducing-sugar assays for both beta-glucanase and xylanase from different fungal origins. Which barley beta-glucan do you recommend for the glucanase assay.

For glucanase assay we recommend the medium viscosity beta-glucan.  We now offer low, medium and high viscosity wheat arabinoxylans, and we think that the low viscosity material will be best (easiest) to use in the reducing-sugar assay.

可吸收碳水化合物检测试剂盒 Available Carbohydrates/Dietary Fiber Assay Kit 货号:K-ACHDF Megazyme中文站

可吸收碳水化合物检测试剂盒

英文名: Available Carbohydrates/Dietary Fiber Assay Kit

货号:K-ACHDF

规格:100 assays of each component

分析物意义:  快速消化和吸收的糖及膳食纤维

Megazyme检测试剂盒优点:检测程序新颖、试剂稳定 

An integrated procedure for the measurement of available carbohydrates and dietary fibre in cereal products, fruit and vegetables and food products. Content:100 assays of each component

An integrated procedure for the measurement of Available
Carbohydrates and Dietary Fiber in cereal products, fruit and
vegetables and food products

Principle (Dietary Fiber):
(α-amylase + amyloglucosidase)
(1) Starch + H2O → glucose

(protease)
(2) Protein + H2O → peptides

(3) Dietary fiber determined gravimetrically following
alcohol precipitation


Principle (Available Carbohydrates):
(sucrase / maltase + β-galactosidase)
(4) Sucrose, maltose and lactose → D-glucose + D-fructose
+ D-galactose


(PGI, hexokinase + glucose-6-phosphate dehydrogenase)
(5) D-Glucose + D-fructose + ATP + NADP+ → gluconate-6-phosphate
+ NADPH + ADP + H+

Kit size: 100 assays of each
Application examples:
Food ingredients, food products and other materials
Method recognition:
Dietary Fibre - AOAC (Methods 985.29, 991.42, 991.43 and 993.19)
and AACC (Methods 32-05.01, 32-07.01 and 32-21.01)

Advantages

  • Very cost effective
  • All reagents stable for > 2 years after preparation
  • High purity / standardised enzymes employed
  • Only kit available
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
  • Simple format

 Q1. There is an issue with the performance of the kit; the results are not as expected.

If you suspect that the Megazyme test kit is not performing as expected such that expected results are not obtained please do the following:

  1. Ensure that you have tested the standard sample that is supplied with the Megazyme test kit.
  2. Send the results of the kit standard, blank samples and the results obtained for your sample,  in the relevant MegaCalc spreadsheet (if available) to Megazyme (cs@megazyme.com). Where available the relevant MegaCalc spreadsheet can be downloaded from where the product appears on the Megazyme website.
  3. State the kit lot number being used (this is found on the outside of the kit box).
  4. State which assay format was used (refer to the relevant page in the kit booklet if necessary).
  5. State exact details of any modifications to the standard procedure that is provided by Megazyme.
  6. State the sample type and describe the sample preparation steps if applicable.

Q2. Should the pH of the sample be adjusted even for samples in acidic media?

The pH of the assay solution after the sample is added should be the same as that of the assay buffer that is supplied with the kit.
Low sample volumes (e.g. 0.1 mL) are not likely to affect the pH of the assay solution and therefore may not require pH adjustment.
Samples above 0.1 mL are more likely to affect the pH of the assay solution and therefore the pH of these samples should be adjusted as described in the data booklet, prior to addition to the assay.

总膳食纤维快速整合试剂盒 Rapid Integrated Total Dietary Fiber Assay Kit 货号:K-RINTDF Megazyme中文站

总膳食纤维快速整合试剂盒

英文名:Rapid Integrated Total Dietary Fiber Assay Kit

货号:K-RINTDF

规格:100 assays per kit

 

This test kit is suitable for the measurement and analysis of Total Dietary Fiber as per Codex Alimentarius definition, updated to be more consistent with in vivo conditions in the human small intestine, i.e. a 4 h incubation time. Under these conditions more accurate measurement of resistant starch is obtained, including phosphate cross-liked starch (RS4). Use of higher enzyme concentrations ensures that resistant maltodextrins produced from non-resistant starch under the incubation conditions of the Integrated Total Dietary Fiber procedure (AOAC Methods 2009.01 and 2011.25 are no longer produced.
In this rapid, improved method, the incubation time with PAA + AMG is reduced to 4 h and the levels of both PAA and AMG are increased to ensure that resistant starch levels obtained with a set of control samples are consistent with ileostomy data. Under these conditions, the DF values obtained for most samples are the same as those obtained with AOAC Methods 2009.01 and 2011.25.
The dietary fiber fractions that are measured with this method are:
1. High Molecular Weight Dietary Fiber (HMWDF) including Insoluble Dietary Fiber (IDF) and Low Molecular Weight Soluble Dietary Fiber (SDFS; soluble dietary fiber which is soluble in the presence of 78% aqueous ethanol), or
2. Insoluble dietary fiber (IDF), Higher Molecular Weight Soluble Dietary Fiber (SDFP; water soluble dietary fiber that precipitates in the presence of 78% aqueous ethanol) and SDFS.
The enzymes used in this method are high purity and effectively devoid of contaminating enzymes active on other dietary fiber components such as β-glucan, pectin and arabinoxylan. They are supplied as freeze-dried powders; allowing the use of glycerol as an internal standard.

For the determination of Total Dietary Fiber in cereal products, foodstuffs, feeds and other materials. 

Principle:
              (Pancreatic α-amylase + amyloglucosidase)
(1) Non-resistant starch + H2O → D-glucose

                      (protease)
(2) Protein + H2O → peptides

(3) IDF (including resistant starch) and alcohol precipitated 
     soluble DF (SDFP) determined gravimetrically

(4) Alcohol soluble DF (SDFS) determined by HPLC

(5) Ash and residual protein determined on DF residues 
     and subtracted

Kit size:                             100 assays
Method:                              Hydrolysis/removal of non-dietary 
                                          fibre components
Total assay time:                ~ 3 h work (over 1-2 days)
Detection limit:                   0.5-100% of sample weight
Application examples: 
Food ingredients, food products and other materials
   

* See McCleary, B. V., Sloane, N & Draga, A. (2015). Determination of total dietary fibre and available carbohydrates: a rapid integrated procedure that simulates in vivo digestion. Starch /Starke, 66, 1-24.

Advantages

  • More rapid measurement – incubation time with PAA + AMG reduced to 4 h in comparison with AOAC 2009.01 (increased levels of enzyme employed)
     
  • DF values for most samples are very similar to those obtained with AOAC Method 2009.01
     
  • Rapid Integrated Total Dietary Fiber method removes all of the limitations that have been identified with AOAC Method 2009.01*
     
  • All reagents stable for > 2 years after preparation
     
  • The method is consistent with the CODEX Alimentarius definition of dietary fiber
     
  • Mega-Calc™ software tool is available from our website for hassle-free raw dataprocessing
     
  • Very competitive price (cost per test)

 

 Q1. How should I prepare the required 78% ethanol solution?

A volume reduction occurs on mixing water with 95% ethanol (or IMS). 1 L of 78% ethanol is prepared by adding 821 mL 95% ethanol (or IMS) to 207 mL H2O.
If using a 1 L volumetric flask, this is best accomplished by placing 821 mL 95% ethanol (or IMS) into the volumetric flask. Dilute to volume with deionised water. Mix well. Check the level and if necessary add more deionised water to bring it back up to the 1 L mark. 

抗性淀粉检测试剂盒 Resistant Starch Assay Kit 货号:K-RSTAR Megazyme中文站

抗性淀粉检测试剂盒

英文名:Resistant Starch Assay Kit

货号:K-RSTAR

规格:100 assays per kit

The Resistant Starch Test kit for the measurement and analysis of resistant starch in plant materials and starch samples.

Colourimetric method for the determination of Resistant Starch
in cereal products and feeds

Principle:
(α-amylase + amyloglucosidase)
(1) Non-resistant starch + H2O → D-glucose + maltose (trace)

(2) Aqueous ethanol wash + centrifugation to remove D-glucose +
maltose

(3) Dissolution of resistant starch pellet in KOH and neutralisation

(α-amylase + amyloglucosidase)
(4) Dissolved resistant starch + H2O → D-glucose

(glucose oxidase)
(5) D-Glucose + H2O + O2 → D-gluconate + H2O2

(peroxidase)
(6) 2H2O2 + p-hydroxybenzoic acid + 4-aminoantipyrine →
quinoneimine + 4H2O

Kit size: 100 assays
Method: Spectrophotometric at 510 nm
Reaction time: ~ 120 min (plus overnight incubation)
Detection limit: 2-100% of sample weight
Application examples:
Plant materials, starch samples and other materials
Method recognition:
AOAC (Method 2002.02), AACC (Method 32-40.01) and CODEX
(Type II Method)

Advantages

  • Very cost effective
  • All reagents stable for > 2 years after preparation
  • Only enzymatic kit available
  • Measures enzyme resistant starch
  • Simple format
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
  • Standard included

 Q1. Should the pH of the sample be adjusted even for samples in acidic media?

The pH of the assay solution after the sample is added should be the same as that of the assay buffer that is supplied with the kit.
Low sample volumes (e.g. 0.1 mL) are not likely to affect the pH of the assay solution and therefore may not require pH adjustment.
Samples above 0.1 mL are more likely to affect the pH of the assay solution and therefore the pH of these samples should be adjusted as described in the data booklet, prior to addition to the assay.

Q2. There is an issue with the performance of the kit; the results are not as expected.

If you suspect that the Megazyme test kit is not performing as expected such that expected results are not obtained please do the following:

  1. Ensure that you have tested the standard sample that is supplied with the Megazyme test kit.
  2. Send the results of the kit standard, blank samples and the results obtained for your sample, in the relevant MegaCalc spreadsheet (if available) to Megazyme (cs@megazyme.com). Where available the relevant MegaCalc spreadsheet can be downloaded from where the product appears on the Megazyme website.
  3. State the kit lot number being used (this is found on the outside of the kit box).
  4. State which assay format was used (refer to the relevant page in the kit booklet if necessary).
  5. State exact details of any modifications to the standard procedure that is provided by Megazyme.
  6. State the sample type and describe the sample preparation steps if applicable.

淀粉总量HK检测试剂盒 Total Starch HK Assay Kit 货号:K-TSHK Megazyme中文站

淀粉总量HK检测试剂盒

英文名: Total Starch HK Assay Kit

货号:K-TSHK

规格:100 assays per kit

A modification of AOAC Method 996.11 AACC Method 76-13.01 RACI Standard Method for the measurement and analysis of total starch in cereal flours and food products. This kit contains an improved α-amylase that allows the amylase incubations to be performed at pH 5.0 (as well as pH 7.0). The method has been further modified by adjusting the D-glucose determination to a hexokinase/glucose-6-phosphate dehydrogenase/NADP+ based format.

UV-method for the determination of Total Starch in grains,
animal feeds, foodstuffs and other materials

Principle:
(α-amylase, 100°C + DMSO)
(1) Starch granules + H2O → maltodextrins

(amyloglucosidase)
(2) Maltodextrins + H2O → D-glucose

(hexokinase)
(3) D-Glucose + ATP → G-6-P + ADP

(glucose-6-phosphate dehydrogenase)
(4) G-6-P + NADP+ → gluconate-6-phosphate + NADPH + H+

Kit size: 100 assays
Method: Spectrophotometric at 340 nm
Total assay time: ~ 90 min
Detection limit: 1-100% of sample weight
Application examples:
Cereal flours, food products and other materials
Method recognition:
AOAC (Method 996.11), AACC (Method 76-13.01), ICC (Standard
Method No. 168), and RACI (Standard Method)

Advantages

  • Very competitive price (cost per test)
  • All reagents stable for > 2 years after preparation
  • Simple format
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
  • Standard included

 

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淀粉总量检测试剂盒 Total Starch (AA/AMG) Assay Kit 货号:K-TSTA-100A Megazyme中文站

淀粉总量检测试剂盒

英文名:Total Starch (AA/AMG) Assay Kit

货号:K-TSTA-100A

规格:100 assays per kit

分析物意义:主要的食品组分

Megazyme检测试剂盒优点:选择用GOPD试剂或己糖激酶或6-磷酸葡萄糖脱氢酶测定D-葡萄糖的快速检测试剂盒

The Total Starch (AA/AMG) test kit is used for the measurement and analysis of total starch in cereal flours and food products. This kit now contains an improved α-amylase that allows the amylase incubations to be performed at pH 5.0 (as well as pH 7.0).

Colourimetric method for the determination of Total Starch in
cereal products, feeds, foodstuffs and other materials

Principle:
(α-amylase, 100°C ± DMSO)
(1) Starch granules + H2O → maltodextrins

(amyloglucosidase)
(2) Maltodextrins + H2O → D-glucose

(glucose oxidase)
(3) D-Glucose + H2O + O2 → D-gluconate + H2O2

(peroxidase)
(4) 2H2O2 + p-hydroxybenzoic acid + 4-aminoantipyrine →
quinoneimine + 4H2O

Kit size: 100 assays
Method: Spectrophotometric at 510 nm
Total assay time: ~ 90 min
Detection limit: 1-100% of sample weight
Application examples:
Cereal flours, food products and other materials
Method recognition:
AOAC (Method 996.11), AACC (Method 76-13.01), ICC (Standard Method
No. 168), and RACI (Standard Method)

Advantages

  • Very competitive price (cost per test)
  • All reagents stable for > 12 months after preparation
  • Simple format
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
  • Standard included

 Q1. Should the pH of the sample be adjusted even for samples in acidic media?

The pH of the assay solution after the sample is added should be the same as that of the assay buffer that is supplied with the kit.
Low sample volumes (e.g. 0.1 mL) are not likely to affect the pH of the assay solution and therefore may not require pH adjustment.
Samples above 0.1 mL are more likely to affect the pH of the assay solution and therefore the pH of these samples should be adjusted as described in the data booklet, prior to addition to the assay.

Q2. Why does the quadruplicate glucose control in your Total Starch Assay Kit have to be incubated?

We feel more comfortable with quadruplicate glucose controls.  If the control is incorrect, or questionable, then all the results are in doubt.

Q3. Why do duplicate samples have to be measured?

Duplicate samples do not have to be measured.  We just suggest this for laboratories starting up. 

Q4. Does the Regular Maize Starch need to be analysed with pre-treating by DMSO? How do you store this Enclosed Control?

The Regular Maize Starch does not require DMSO pre-treatment.  The value should be about 84% with a moisture content of about 12%, the final dry weight value is about 96-97%.  Store the sample at room temperature, dry.

Q5. There is an issue with the performance of the kit; the results are not as expected.

If you suspect that the Megazyme test kit is not performing as expected such that expected results are not obtained please do the following:

  1. Ensure that you have tested the standard sample that is supplied with the Megazyme test kit.
  2. Send the results of the kit standard, blank samples and the results obtained for your sample, in the relevant MegaCalc spreadsheet (if available) to Megazyme (cs@megazyme.com). Where available the relevant MegaCalc spreadsheet can be downloaded from where the product appears on the Megazyme website.
  3. State the kit lot number being used (this is found on the outside of the kit box).
  4. State which assay format was used (refer to the relevant page in the kit booklet if necessary).
  5. State exact details of any modifications to the standard procedure that is provided by Megazyme.
  6. State the sample type and describe the sample preparation steps if applicable.

Q6. Does your kit with DMSO solubilise starch that has been vitrified due to malting/kilning?

Yes.  We believe that the DMSO step will solubilise vitrified starch in malt.  Make sure that the malt is milled to pass a 0.5 mm screen.  You could vary the time of cooking with DMSO to check solubilisation (i.e. 5 minutes, 10 minutes, or even up to 1 hour).

Q7. Is it possible to differentiate between gelatinised and ungelatinised starch in finished products, such as dog food, using the Total Starch Kit?

We think that there is a better chance of success using the Megazyme Starch Damage Kit.

Q8. Is it possible to use the Total Starch Kit to measure starch levels in plasterboard and related products?

There should be no problem in measuring the starch in plasterboard.  I suggest that you grind about 100 g in a kitchen blender and then fine mill to pass 0.5 mm screen. Run a standard assay, but adjust volume to 10 mL after alpha-amylase treatment. Keep a close check on the pH.  Plasterboard may push the pH value up (pH up to about 8 should be fine).  You may be advised to run a DMSO format concurrently just to be sure.  When you treat with amyloglucosidase, I would advise that you take 0.2 and 0.4 mL aliquots of digest (to get the colour up), also, be careful about checking the pH.

Q9. Can the Total Starch Kit be used for samples containing 20% fat or higher?

A 20% fat content could cause a problem for the method.  We suggest that the sample be defatted before analysis for starch.

Q10. Do you have any kit or procedures for the determination of extractable starch in corn? This is of particular concern in the corn wet milling. I presume that extractable starch in corn is not the same as total starch?

Our Total Starch Assay Kit could measure starch left in a residue, or starch extracted. No method could measure potential extractable starch, as this will depend on numerous factors, including processing equipment, conditions etc.

Q11. Can I use another buffer instead of the MOPS with your Total Starch Assay Kit? If so, which would be suitable and easily prepared from commonly available laboratory reagents?

You can use phosphate buffer at the same concentration.

Q12. I wish to measure Total Starch in several products. These products contain 10-20% starch + maltodextrins at similar levels. Is it possible to remove the maltodextrins from the sample? Will ethanol work?

Most of the maltodextrins can be removed with 50% ethanol washing.  If the starch is not gelatinised, it can be washed with cold water.  This will remove all of the soluble maltodextrins, but the starch will spin down.  If the starch has been gelatinised, then the best material which can be used for washing is 50% ethanol.

Q13. Could your Total Starch Assay Kit (K-TSTA) be used with success to measure total starch in plant tissues (samples of roots and shoots of maple trees)?

Yes, the Total Starch Kit can be used to measure starch in roots and shoots etc. 

Q14. Is the accuracy of the Total Starch test affected by the presence of other inorganic chemicals and ground calcium carbonate in pulp?

We think that calcium carbonate etc. will not cause any problems.  However, this of course depends on the amount present and if it changes the pH of the incubation mixture.

Q15. Does the Megazyme Total Starch method work well on all the new chemically modified starches that are now appearing, e.g. highly crosslinked, dextrinised and highly propylene oxide substituted?

The method will work for some chemically modified starches (e.g. crosslinked) however, if the degree of chemical modification is high, there will be an underestimation as the modification will interfere with complete hydrolysis to glucose and subsequent measurement.

Q16. Is it necessary to pre-wash ground cereal samples prior to analysis for Total Starch?

You only need to wash samples which you feel may contain glucose and/or maltodextrins, e.g. breakfast cereals.  There is little glucose in ground cereals, so it is not necessary to pre-wash these materials.

Q17. What is the stability of the enzymes from the Total Starch Kit?

The enzymes from this kit are stable at room temperature for at least 6 months.  At 4˚C, they are stable for several years.

Q18. Can the Total Starch Kit determine the degree of gelatinisation? Sample : Corn Flour.

The Starch Damage Kit may be best for this.  If the starch is gelatinised and dried before analysis the correct results for gelatinisation will not be obtained.

Q19. Are there any limits to the sensitivity of the Total Starch Kit?

The Total Starch Kit can accurately measure starch levels as low as 1% w/w.

Q20. What is the sensitivity and how much is the absorbance of glucose standard (100 micrograms)?

The absorbance for 100 micrograms of glucose (in 3 mL of GOPOD Reagent) is about 0.97.

Q21. Is it possible to raise sensitivity by modifying dilution of GOPOD reagent?

Yes, you can reduce the volume of GOPOD to 1 mL and use micro cuvettes.  This will increase sensitivity by ~ 3-fold.

Q22. Does DMSO solubilise resistant starch, i.e. crystallised amylose and amylopectin?

DMSO does solubilise resistant starch (crystallise amylose and amylopectin).  The only starch material we have had problems in dissolving in DMSO is potato amylose.

Q23. When analysing samples containing sugars, an 80% v/v solution of ethanol is used to solubilise and remove the sugars. About how large are the smallest dextrins that are left in the starch (not solubilised) in this treatment?

We believe that for starch fragments, oligosaccharides of a DP up to 10 would be soluble in 80% alcohol.  The degree of solubility of other oligosaccharides would depend on the sugar type and linkage type.

Q24. What is the sensitivity of the Total Starch Method for measurement in liquids containing low levels of starch?

The Total Starch Kit can be used for liquids containing as little as 200 micrograms per mL with some adjustments of conditions, as below:
Mix 0.5 mL of sample with 0.5 mL of 100 mM sodium acetate buffer (pH 4.5). Incubate at 40˚C and add 0.1 mL of Amyloglucosidase and incubate for 30 minutes.  Add GOPOD reagent as usual.  You will need to run an AMG blank as this enzyme preparation contains a very small amount of glucose.

Q25. AMYLOGLUCOSIDASE. The activity is stated as being 3260 U/mL (Soluble Starch). How was this determined?

The AMG activity was determined with soluble starch as substrate (10 mg/mL) in 0.1 M sodium acetate buffer at pH 4.5 and 40˚C.  One Unit is the amount of enzyme required to hydrolyse one micromole of maltose per minute (i.e. to release 2 micromoles of glucose).  Glucose release is measured with Glucose Determination Reagent.

Q26. I wish to know if it is possible to perform the assay under acidic conditions? I also need to alter the pH of the MOPS/amylase mixture to pH 3 or 4. Is it known if the amylase supplied with the Total Starch Assay Kit has activity at such a low pH?

We can assure you that the Total Starch Kit will not work if incubations with the thermostable alpha-amylase are performed at pH 3 or 4.  This enzyme is inactivated at pH values below 5.0.  You may wish to look at a method using just amyloglucosidase which is quite active down to pH 4.0.  Check the old AOAC procedure for starch.

Q27. We are currently using your kits for Total Starch, Starch Damage and Sucrose/Glucose/Lactose Assay procedures. How accurate are the volumes contained? Can they be diluted entirely, to avoid wasting the contents while transferring?

When we dispense the enzymes we usually include an extra 5% in each vial, so yes, you can dilute the whole vial.  When you do this, please divide into aliquots and store them frozen.

Q28. Which analytical procedure would you recommend for determination of total starch residues in brewing wort and final beer. Megazyme’s procedure – AA/AMG’97, only refers to solid (and not solubilised) starch.

You can use the standard procedure. We would recommend that you treat 2 mL of beer or wort with 8 mL of ethanol, stir and centrifuge (3,000 rpm).  Wash the pellet with 10 mL of 80% ethanol.  Then dissolve/suspend the pellet in 2 mL of sodium acetate buffer (pH 4.5, 0.1 M) and cook at 100˚C for 10 min.  Then add 0.1 mL AMG from the Total Starch Kit and proceed according to the method.  You will have to determine the degree of dilution for yourself.  Treat 0.1 mL with GOPOD etc.