CN-121975906-A - Method for detecting mould in feed by ATP method

Abstract

The invention relates to a method for detecting mould in feed by an ATP method, which comprises the steps of S1, sequentially carrying out first mixing and extraction treatment on a feed sample and normal saline, collecting supernatant as an extracting solution, S2, sequentially carrying out second mixing and cracking treatment on the extracting solution and a benzalkonium bromide solution with the concentration of 0.10-0.20% to obtain a cracking solution, S3, carrying out third mixing on the cracking solution and an ATP reaction reagent to obtain a liquid to be detected, and S4, measuring the RLU value of the liquid to be detected by using a handheld ATP luminometer. The method has the advantages of high detection speed and high accuracy, and can detect and control the mildew of the feed in time in early stage of the mildew of the feed.

Inventors

• LI YUXUAN
• ZHANG YAMENG
• CAO DIE
• LI FENG
• GU ZHIGUANG

Assignees

• 蓝星安迪苏南京有限公司

Dates

Publication Date

20260505

Application Date

20251229

Claims (10)

1. 1. A method for detecting mold in feed by an ATP method, the method comprising: S1, sequentially carrying out first mixing and extraction treatment on a feed sample and normal saline, and collecting supernatant as an extracting solution; s2, sequentially carrying out second mixing and cracking treatment on the extracting solution and the benzalkonium bromide solution with the concentration of 0.10-0.20% to obtain a cracking solution; S3, carrying out third mixing on the lysate and the ATP reaction reagent to obtain a liquid to be detected; S4, measuring the RLU value of the liquid to be measured by using a handheld ATP luminometer.
2. 2. The method according to claim 1, wherein in step S1, the ratio of the weight of the feed sample to the volume of the physiological saline is 1:7-10.
3. 3. The method of claim 2, wherein the first mixing comprises vortexing at 1000-1500rpm for 30-60s; the extraction process comprises standing for 3-8min.
4. 4. The method according to claim 1, wherein in step S2, the volume ratio of the extract to the benzalkonium bromide solution is 1:1-2.
5. 5. The method of claim 4, wherein the second mixing comprises repeatedly blowing and sucking 5-10 times using a pipette; The lysis treatment comprises incubation for 3-15min.
6. 6. The method according to claim 1, wherein in step S3, the volume ratio of the lysate to the ATP-reactive reagent is 1:1-2; The ATP reagent comprises a substrate containing luciferases and phosphate buffer.
7. 7. The method of claim 6, wherein the third mixing comprises repeatedly pipetting 5-10 times using a pipette.
8. 8. The method of claim 1, wherein the RLU value of the test solution is determined using a handheld ATP luminometer within 5 minutes of the test solution preparation.
9. 9. The method of claim 1, wherein if the RLU value is >50, it is indicative of the presence of mold in the feed sample.
10. 10. The method of claim 9, wherein the mold comprises Xie Wali a aspergillus, fusarium verticillatum, and penicillium oxalicum.

Description

Method for detecting mould in feed by ATP method Technical Field The disclosure relates to the technical field of mold detection, in particular to a method for detecting mold in feed by an ATP method. Background The formula feed and the feed raw materials are often mildewed due to environmental factors such as storage conditions, especially when the cereal feed raw materials are stored, the mildewed cereal feed raw materials are often mildewed due to the rise of air temperature and humidity, mycotoxins produced by the metabolism of the mildewed cereal feed are extremely harmful to the health of people and livestock, so that the mildewed feed is not edible, and huge losses are caused to crop economy and livestock breeding. Therefore, the units or individuals such as farms or feed suppliers need to detect, prevent and control the mildew phenomenon, and especially the timely detection and prevention and control of the mildew phenomenon in early stage of feed mildew are significant. The current common method for evaluating mildew of feed in the feed and livestock industry mainly comprises ① sensory evaluation, wherein the method is used for judging whether mildew is generated or not on sense of smell by relying on experience, whether feed or feed raw materials are agglomerated or not, whether mildew spots, mildew spots or hyphae are generated or not is visually used as a judging standard, the method is low in evaluation sensitivity, a non-long-term practitioner is difficult to accurately identify the visual state of mildew smell and hyphae growth of the feed, early mildew of the visible hyphae is easy to miss-check, and the method (GB/T13092) for counting ② total moulds is required to be observed after carefully sampling and culturing on a specific plate culture medium for 2-3 days, but generates larger errors on the final colony number due to sampling accuracy and inoculation accuracy, and is time-consuming in mildew detection due to longer waiting time, and ③ carbon dioxide monitoring method is based on the total quantity of carbon dioxide generated by metabolism in a detection system and corresponds to the total quantity of moulds, but is often error due to instability, and the method is inconvenient for on site operation due to special falling equipment. At present, the prior art cannot simultaneously ensure the detection speed and the accuracy, and is difficult to prevent and control the mildew of the feed, so that a method with high detection speed, real-time detection and higher accuracy is needed urgently so as to timely detect and prevent and control the mildew of the feed in early stage of the mildew of the feed. Disclosure of Invention The method is high in detection speed and accuracy, and can detect and control the mildew of the feed in time in early stage of the mildew of the feed. In order to achieve the above object, the present disclosure provides a method for detecting mold in feed by an ATP method, the method comprising: S1, sequentially carrying out first mixing and extraction treatment on a feed sample and normal saline, and collecting supernatant as an extracting solution; s2, sequentially carrying out second mixing and cracking treatment on the extracting solution and the benzalkonium bromide solution with the concentration of 0.1-0.2% to obtain a cracking solution; S3, carrying out third mixing on the lysate and the ATP reaction reagent to obtain a liquid to be detected; S4, measuring the RLU value of the liquid to be measured by using a handheld ATP luminometer. Optionally, in step S1, the ratio of the weight of the feed sample to the volume of the physiological saline is 1:7-10. Optionally, the first mixing comprises vortexing at 1000-1500rpm for 30-60s; the extraction process comprises standing for 3-8min. Optionally, in step S2, the volume ratio of the extracting solution to the benzalkonium bromide solution is 1:1-2. Optionally, the second mixing comprises repeatedly blowing and sucking 5-10 times by using a pipette; The lysis treatment comprises incubation for 3-15min. Optionally, in step S3, the volume ratio of the lysate to the ATP reagent is 1:1-2; The ATP reagent comprises a substrate containing luciferases and phosphate buffer. Optionally, the third mixing comprises repeatedly blowing and sucking 5-10 times by using a pipette. Optionally, the RLU value of the test solution is determined using a handheld ATP luminometer within 5min of the test solution preparation. Alternatively, if the RLU value is >50, this represents the presence of mould in the feed sample. Alternatively, the mold comprises Xie Wali A. Oryzae, fusarium verticillatum, and Penicillium oxalicum. Through the technical scheme, the disclosure provides a method for detecting mould in feed by using an ATP method, which has the following advantages: (1) The method is simple, the detection speed is high, the detection can be completed within 30 minutes, and the timeliness of the mildew detection of the feed is remark