Method for losslessly and accurately evaluating alimentary canal emptying time of haliotis discus hannai

Abstract

The invention discloses a method for losslessly and accurately evaluating alimentary canal emptying time of haliotis discus hannai, and belongs to the technical field of aquaculture. After the haliotis discus hannai is fed with artificial compound feed containing chromium sesquioxide (Cr2O3), the defecation amount of the haliotis discus hannai is measured every 2-4 h, the alimentary canal emptying characteristic of the haliotis discus hannai is calculated by calculating the content of Cr2O3 in excrement, and the time spent when the proportion of the accumulated Cr2O3 amount reaches or exceeds 99% of the acceptable daily intake of the haliotis discus hannai is defined as the alimentary canal emptying time of the haliotis discus hannai. The alimentary canal emptying time of the haliotis discus hannai is accurately evaluated through a lossless and non-invasive method, and an important reference is provided for formulating a scientific and reasonable feeding strategy of the haliotis discus hannai and improving the breeding efficiency.

Description

Technical Field

[0001] This invention belongs to the field of abalone farming technology, specifically relating to a non-destructive and accurate method for measuring the emptying time of the digestive tract of wrinkled abalone. Background Technology

[0002] Abalone is one of the world's most valuable fishery products. Scientific and reasonable feeding strategies have a positive impact on its aquaculture. Establishing a digestive tract emptying model for this species and understanding its digestive tract emptying time are essential for formulating its feeding strategies.

[0003] Determining the gastrointestinal emptying time of aquatic animals is crucial for developing feeding strategies for that species, and this data is often obtained through dissection. However, given the delicate and fragile digestive tract of abalone, and the adhesion between the stomach wall and the hepatopancreas, it is difficult to completely dissect the digestive tract and obtain sufficient contents. Therefore, data obtained using dissection methods often contain significant biases. Thus, establishing a precise, non-destructive, and non-invasive method for determining the gastrointestinal emptying time of the wrinkled abalone is of great importance for developing a scientifically sound feeding strategy for this species. Summary of the Invention

[0004] To address the problem that the gastrointestinal emptying time of abalone is difficult to accurately measure through dissection, this invention provides a non-destructive and accurate method for evaluating the gastrointestinal emptying time of abalone. This method can accurately measure the gastrointestinal emptying time of abalone without causing harm, and can provide an important reference for formulating feeding strategies for abalone.

[0005] To achieve the above objectives, the present invention adopts the following technical solution: A non-destructive and accurate method for evaluating the gastrointestinal emptying time of abalone is proposed. After feeding abalone with an artificial compound feed containing 1.0-2.0% Cr2O3, the amount of feces is measured every 2-4 hours. The gastrointestinal emptying characteristics are estimated by calculating the Cr2O3 content in the feces. The time taken when the cumulative amount of Cr2O3 excreted reaches or exceeds 99% of the ingested amount is defined as the gastrointestinal emptying time.

[0006] Abalone were fed artificial formulated feed daily from 16:00 to 18:00 for more than 5 days. Then, at the same time, they were fed a formulated feed containing Cr2O3 for only one day. From the next day, they were fed a formulated feed without Cr2O3. Two to four hours after feeding the Cr2O3-containing feed (18:00-20:00 on the same day), the uneaten feed was removed with a sieve and weighed. The amount of formulated feed / Cr2O3 ingested by the abalone was calculated based on the feed's solubility index. After feeding the Cr2O3-containing feed, abalone feces were collected every 2-4 hours with a sieve (0.02-0.04 mm aperture) and all water was changed. The feces were then freeze-dried, weighed, and the Cr2O3 content was determined. The Cr2O3 content in each feces collection was recorded. Collection was stopped when the cumulative Cr2O3 in the feces reached 99% of the ingested amount. The time from feeding to this point is the abalone's digestive tract emptying time.

[0007] Preferably, the abalone artificial compound feed is a general commercial compound feed, and the artificial compound feed with added Cr2O3 contains 1.5% Cr2O3 (except for the addition of Cr2O3, the formula is the same as that of ordinary compound feed).

[0008] As a preferred option, abalone should be fed an artificial compound feed containing or without Cr2O3 at 5% of their wet body weight.

[0009] Preferably, the intake of Cr2O3 is estimated by the amount of compound feed containing Cr2O3 ingested; the excretion of Cr2O3 is estimated by measuring the chromium ion content in the fecal solution.

[0010] Preferably, the compound feed containing Cr2O3 is a flake-shaped compound feed.

[0011] Preferably, the time for collecting feces after feeding compound feed containing Cr2O3 is no more than 72 hours.

[0012] This method can also be tried to be applied to other farmed abalone species besides the wrinkled abalone.

[0013] The beneficial effects of this invention are: Based on the characteristic that Cr2O3 is not digested and absorbed by abalone and has no negative impact on abalone, this invention uses Cr2O3 as an indicator in artificial compound feed to construct a non-destructive and accurate technology for measuring the gastrointestinal emptying time of wrinkled abalone. This technology achieves accurate measurement of gastrointestinal emptying time without causing secondary damage to the organism and has great application prospects. Detailed Implementation

[0014] The present invention will be further described below with reference to specific implementation methods.

[0015] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below. The following non-limiting embodiments can enable those skilled in the art to more fully understand the present invention, but do not limit the present invention in any way.

[0016] Example 1 Select abalone with a shell length of 6.89±0.36 cm and a wet weight of 37.33±5.86 g. Discus fish Hannah Thirty animals were placed in six separate water tanks (45 cm × 30 cm × 30 cm). The seawater temperature was 18.7 ± 0.5℃, pH = 8.3, salinity 30 ± 1, and dissolved oxygen concentration > 6 mg / L.

[0017] All abalone were fed a standard flaky feed (crude ash 10.70±0.57%, moisture 6.44±0.04%, crude protein 40.40±0.50%, crude fat 3.26±0.23%, crude carbohydrates 39.20%; 20mm×30mm×3mm) for 15 days at 5% of their wet body weight to ensure they reached satiation. Feeding was done daily at 16:00, and uneaten feed was collected and the seawater replaced at 8:00 the following morning. After the abalone adapted to the flaky feed, a fixed amount (weighed using an analytical balance with an accuracy of 0.0001 g) of flaky feed containing 1.5% Cr2O3 (the formula was the same as the standard flaky feed except for the added Cr2O3) was given for one day, after which the standard flaky feed was continued. During the 72 hours following the start of the Cr2O3-containing flaky feed, uneaten feed and feces were collected every four hours, and the seawater was replaced.

[0018] At each sampling time point, uneaten food was first removed using a sieve and placed into centrifuge tubes, which were then stored at -20°C. A suction device was then used to create a water flow to flush away any feces that might be adhering to the abalone's body surface. All seawater, along with the feces, was then poured out. The remaining feces were filtered through a pre-weighed 0.04 mm mesh sieve and stored at -20°C. The sampling time at each time point was controlled to be within 10 minutes to minimize stimulation and disturbance to the abalone.

[0019] The collected uneaten feed was dried at 110℃ for 12 hours and weighed. Using seawater identical to the aquaculture environment, flake feed was placed in the water and soaked for 16 hours or 4 hours, the same amount of feed consumed. This soaking time was then used to calculate the feed loss rate to correct for apparent feed intake. Apparent feed intake was calculated as follows: The total amount of feed fed to each tank was reduced by the weight of uneaten feed and feed lost due to leaching; Actual intake (g) = Feeding amount (g) - [Uneaten feed dry weight (g) + Loss (g)]; where loss = Loss rate of feed dry weight after soaking × Feeding amount; Loss rate (%) = [(Feed weight before soaking - Feed weight after soaking) / Feed weight before soaking] × 100.

[0020] Accurately weigh 0.01–0.02 g of freeze-dried feces and place it in a 55 mL microwave digestion vessel. Add 8 mL of 65% concentrated nitric acid (CNW, guaranteed reagent) and 2 mL of 30% hydrogen peroxide solution (Xilong Scientific, analytical reagent) for 24 hours of pre-digestion. Tighten the lid of the microwave digestion vessel and place it in a microwave digestion system (CEM MARS, Xpress). Digest at 1.2 kW power at 160°C for 30 min, then at 200°C for 20 min to completely oxidize chromium trioxide in the feces to water-soluble dichromate. Dilute the digestion solution to 45 mL with 1% nitric acid. Filter all the digestion solution using a 0.22 μm pore size disposable syringe filter (Millipore, SLGP033RB) and bring the volume to 50 mL. The filtrate was then diluted 10-fold with 3% nitric acid, and the chromium content in the diluted solution was determined using inductively coupled plasma atomic emission spectrometry (ICP-Agilent, 5110VDV). Standard curves were plotted using solutions with chromium concentrations of 0.008, 0.04, 0.1, 0.2, 0.3, 0.5, 0.8, and 1 μg / ml, prepared according to environmental calibration standards (Agilent, 5183-4688). The operating parameters of the ICP-AES were: plasma gas flow rate 12 L / min, auxiliary gas flow rate 1 L / min, nebulizer gas flow rate 0.8 L / min, RF power 1.2 kW, 99.996% argon, 3 repetitions, and wavelengths of 267.716 nm and 205.560 nm. The formula for calculating the Cr2O3 content in the feces excreted by abalone at different time periods is: Cr2O3 content in feces (%) = Chromium content in diluent (mg / mL) × Dilution factor × Volume of final volume (mL) / Dry weight of feces to be digested (mg) × 100%.

[0021] The amount of chromium trioxide excreted during a given period is calculated by multiplying the total amount of feces excreted over a specific time period by the Cr2O3 content in the feces. The evacuation percentage at a given time point from the ingestion of the labeled feed is measured as the percentage of the total Cr2O3 excreted in all previous time periods relative to the total ingested Cr2O3. The total time it takes for the labeled feed to be emptied is the total time from the start of labeled feed ingestion until the evacuation percentage reaches 99% or higher.

[0022] The percentage of Cr2O3 excreted by the abalone at different time points was summed up. In the first 12 hours after the abalone ingested the marked feed, the excretion / ingestion ratio was 8.496%, which increased to 75.168% between 12 and 24 hours; it reached 98.215% at 40 hours and 99.428% at 44 hours. The 99% threshold was reached between 40 and 44 hours.

[0023] Example 2 Western-style abalone of similar specifications ( Giant halibut Replace the wrinkled abalone in Example 1, and the rest of the processing mode is the same as in Example 1.

[0024] The percentage of Cr2O3 excreted by abalone at different time points was summed up. In the first 12 hours after ingestion of the labeled feed, the excretion / ingestion ratio was 8.930%, rising to 83.459% between 12 and 24 hours. The ratio reached 99% between 48 and 52 hours, specifically 98.850% at 48 hours and 99.243% at 52 hours.

[0025] Example 3 Hybrid abalone of similar size ( Haliotis discus hannai ♀ × Haliotis fulgens ♂ Replace the wrinkled abalone in Example 1, and the rest of the processing mode is the same as in Example 1.

[0026] The percentage of Cr2O3 excreted by hybrid abalone at different time points was summed up. In the first 12 hours after ingestion of the labeled feed, the excretion / ingestion ratio was 9.737%, rising to 78.528% between 12 and 24 hours. The percentage exceeding 99% was reached between 40 hours (98.619%) and 44 hours (99.235%).

Claims

1. A non-destructive and accurate method for measuring the gastrointestinal emptying time of wrinkled abalone, characterized in that: After feeding wrinkled abalone with artificial compound feed containing Cr2O3, their defecation volume was measured every 2-4 hours. The digestive tract emptying characteristics were estimated by calculating the Cr2O3 content in the feces. The time taken when the cumulative amount of Cr2O3 excreted reached or exceeded 99% of the intake was defined as the digestive tract emptying time.

2. The method according to claim 1, characterized in that: The method specifically includes the following steps: (1) Feed abalone with a compound feed that does not contain Cr2O3 for more than 5 consecutive days from 16:00 to 18:00 every day. Then feed them with a compound feed containing Cr2O3 at the same time for only 1 day. From the next day, resume feeding them with a compound feed that does not contain Cr2O3. (2) After feeding the abalone with a compound feed containing Cr2O3 for 2-4 hours, remove the remaining feed with a sieve and weigh it. Then, calculate the amount of compound feed and Cr2O3 ingested by the abalone based on the feed loss rate. (3) After feeding the abalone with a compound feed containing Cr2O3, collect the abalone feces with a sieve and change all the water every 2-4 hours. Then freeze-dry and weigh the feces, and determine the Cr2O3 content. (4) Record the Cr2O3 content in the feces each time it is collected. Stop collecting when the cumulative amount of Cr2O3 in the feces reaches 99% of its intake. The time from feeding to this point is the abalone's digestive tract emptying time.

3. The method according to claim 2, characterized in that: The Cr2O3 content in the abalone artificial compound feed is 1.0-2.0 wt%.

4. The method according to claim 2, characterized in that: Feed the abalone with a compound feed containing Cr2O3 or a compound feed without Cr2O3 at 5% of their wet body weight.

5. The method according to claim 2, characterized in that: The intake of Cr2O3 was estimated by the amount of compound feed containing Cr2O3 ingested; the excretion of Cr2O3 was estimated by measuring the chromium ion content in the fecal solution.

6. The method according to claim 2, characterized in that: The screen mesh size is 0.02-0.04 mm.

7. The method according to any one of claims 1-6, characterized in that: It can be applied to other cultured gastropod species besides the wrinkled abalone.