Pichia kudriavzevii FJM18 and application thereof in mycoprotein feed

Abstract

The invention relates to the technical field of microorganisms, and discloses pichia kudriavzevii FJM18 and an application of the pichia kudriavzevii FJM18 in a mycoprotein feed, and the preservation number of the pichia kudriavzevii FJM18 is CGMCC (China General Microbiological Culture Collection Center) No.30405. The FJM18 yeast can effectively convert a substrate into feed rich in nutrition, the nutritional value of the feed is improved, meanwhile, emission of organic pollutants is reduced, and environmental protection is facilitated. Besides, the application of the FJM18 yeast in preparation of a biological feed additive and a biological feed can also improve the biological activity of the feed, enhance the immunity of the poultry and promote the healthy growth of the poultry.

Description

Technical Field

[0001] The present invention relates to the field of microbial technology, and in particular to Pichia kudriavzevii FJM18 and application thereof in bacterial protein feed. Background Art

[0002] As a high-quality feed resource, bacterial protein feed is gradually gaining more and more attention and recognition in the breeding industry. Bacterial protein feed not only has rich nutrients, such as protein, amino acids, minerals, etc., but its unique bioactive substances can also promote animal digestion and absorption, and improve breeding efficiency.

[0003] Compared with traditional feed, bacterial protein feed has many advantages. First, bacterial protein feed has high protein content, excellent quality, and is easy to digest and absorb. Secondly, bacterial protein feed is rich in a variety of beneficial microorganisms and enzymes, which can improve the intestinal microecological environment of animals and enhance the immunity of animals. In addition, bacterial protein feed also has good environmental performance, which can effectively reduce the discharge of breeding waste and alleviate the pressure of environmental pollution.

[0004] In the aquaculture industry, bacterial protein feed has a wide range of applications. It can be used as a feed additive for pigs, chickens, cattle, sheep and other animals, and can also be used as a feed raw material for aquatic animals. By using bacterial protein feed, not only can the growth rate and feed utilization rate of animals be increased, but also the quality of animal products can be improved, and the overall economic benefits of the aquaculture industry can be improved.

[0005] Therefore, developing a new bacterial protein feed can better meet the needs of the breeding industry and improve animal production performance and product quality. Summary of the invention

[0006] In view of this, the present invention proposes a Pichia kudriavzevii FJM18 and an application thereof in bacterial protein feed, aiming to better meet the needs of the breeding industry.

[0007] The invention provides a Pichia kudriavzevii FJM18, and the preservation number of the Pichia kudriavzevii FJM18 is CGMCC No.30405.

[0008] The present invention also proposes an application of the Pichia kudriavzevii FJM18, which includes at least one of the following:

[0009] A) application of the Pichia kudriavzevii FJM18 in fermenting wheat bran;

[0010] B) application of the Pichia kudriavzevii FJM18 in fermentation of distiller's grains;

[0011] C) Application of the Pichia kudriavzevii FJM18 in the preparation of biological feed additives;

[0012] D) Application of the Pichia kudriavzevii FJM18 in preparing biological feed;

[0013] E) Application of the Pichia kudriavzevii FJM18 in degrading organic pollutants;

[0014] F) Application of the Pichia kudriavzevii FJM18 in improving the apparent metabolizable energy of poultry.

[0015] Preferably, the organic pollutant is wheat bran or distiller's grains.

[0016] The present invention also provides a bacterial agent, which comprises the Pichia kudriavzevii FJM18.

[0017] The present invention also provides a method for preparing bacterial protein feed, comprising:

[0018] The fermentation substrate is fermented by using the Pichia kudriavzevii FJM18 to obtain the bacterial protein feed.

[0019] Preferably, the fermentation substrate is wine lees and ammonia water.

[0020] Preferably, the fermentation substrate is wheat bran, urea and ammonium sulfate.

[0021] Compared with the prior art, the present invention has the following beneficial effects:

[0022] The Pichia kudriavzevii FJM18 of the present invention has significant application value in multiple fields. In the process of fermenting wheat bran and distiller's grains, the FJM18 yeast can effectively convert the substrate into nutrient-rich feed, improve the nutritional value of the feed, and reduce the emission of organic pollutants, which is beneficial to environmental protection. In addition, the application of the FJM18 yeast in the preparation of biological feed additives and biological feed can also improve the biological activity of the feed, enhance the immunity of poultry, and promote the healthy growth of poultry.

[0023] At the same time, the FJM18 yeast of the present invention also has significant advantages in degrading organic pollutants. By using FJM18 yeast to ferment organic pollutants such as wheat bran or distiller's grains, the organic pollutants can be effectively converted into useful feed, achieving waste resource utilization and reducing environmental pollution.

[0024] In addition, the FJM18 yeast of the present invention also has significant applications in improving the apparent metabolic energy of poultry. By adding FJM18 yeast to poultry feed, the metabolic energy of poultry can be effectively improved, the digestive capacity of poultry can be enhanced, the utilization rate of feed can be improved, the feed cost can be reduced, and the production efficiency of poultry can be improved.

[0025] The present invention also provides a method for preparing bacterial protein feed using FJM18 yeast, which has the advantages of being simple, easy to operate, low cost, etc., and is suitable for large-scale production and application. Through the preparation method of the present invention, nutrient-rich bacterial protein feed can be obtained, providing a strong guarantee for the healthy growth of poultry.

[0026] In summary, the Pichia kudriavzevii FJM18 of the present invention has broad application prospects and important practical application value, and is expected to make important contributions to the sustainable development of animal husbandry. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Various other advantages and benefits will become apparent to those of ordinary skill in the art by reading the detailed description of the preferred embodiments below. The accompanying drawings are only for the purpose of illustrating the preferred embodiments and are not to be considered as limiting the present invention. Moreover, the same reference symbols are used throughout the accompanying drawings to represent the same components. In the accompanying drawings:

[0028] Figure 1 The growth curves of FJM18 and FJM60 of the present invention in basic peptone medium and basic ammonia water medium, respectively (OD value is measured every 2 hours);

[0029] Figure 2 These are the growth curves of FJM18 and FJM60 of the present invention when different seed solutions were transferred to different fermentation medium (J is a basic peptone medium, and N is a basic ammonia water medium) (OD value was measured every 4 hours). DETAILED DESCRIPTION

[0030] Exemplary embodiments of the present disclosure will be described in more detail below with reference to the accompanying drawings. Although exemplary embodiments of the present disclosure are shown in the accompanying drawings, it should be understood that the present disclosure can be implemented in various forms and should not be limited by the embodiments described herein. On the contrary, these embodiments are provided in order to enable a more thorough understanding of the present disclosure and to fully convey the scope of the present disclosure to those skilled in the art. It should be noted that, in the absence of conflict, the embodiments of the present invention and the features described in the embodiments can be combined with each other. The present invention will be described in detail below with reference to the accompanying drawings and in combination with the embodiments.

[0031] Example 1 Isolation and screening of yeast

[0032] 1. Initial screening

[0033] Five samples of naturally fermented feed and three samples of chicken intestinal contents (including jejunum, ileum and cecum) were collected. The samples were screened and sequenced using gradient dilution and selective culture medium technology. In view of the positive correlation between the number of viable bacteria and bacterial protein, we adopted a simple primary screening method. The nitrogen sources such as peptone in the screening culture medium YPD were replaced with ammonium sulfate, ammonium chloride, urea and ammonia water to prepare a solid culture medium. Subsequently, a single colony was selected and inoculated into YPD liquid culture medium and cultured at 30°C and 180rpm for 15 hours. Afterwards, the bacterial solution was diluted to -5 and poured into the plate using the 0.5mL pouring method. After 2-3 days of culture, the colonies were counted.

[0034] A total of 15 yeast strains were isolated in the preliminary screening (see Table 1). Among these 15 strains and 3 laboratory-preserved strains, we found that strain FJM16 had the highest viable count, followed by FJM15 and CJM21 (see Table 2 for the viable count results of colony morphology, unit: log (cfu / mL)). In addition, 5 strains had large colonies, among which FJM18 had the highest viable count (see Table 3 for the viable count results of large colonies in colony morphology, unit: log (cfu / mL)). Strain FJM18 is Pichia kudr iavzev ii, which was deposited in the General Microbiology Center of the China Microbiological Culture Collection Administration on April 22, 2024, with the deposit number CGMCC No.30405.

[0035] Table 1

[0036]

[0037]

[0038] Table 2

[0039] Species name YPD Ammonium sulfate Ammonium chloride Urea SC19-1 7.660865 7.534026 7.305351 7.421604 SC16-1 7.764923 7.838219 7.781037 7.726727 Pichia stipitis 7.217484 6.986772 6.919078 6.799341 FJM12 7.838849 7.740363 7.745855 7.869232 FJM13 7.866878 3 7.751279 7.812913 FJM15 7.921166 7.905796 7.964731 7.938019 FJM16 8.255273 8.259355 8.20412 8.20412 CJM21 7.867467 7.799341 7.85187 7.752048 CJM26 7.658965 7.644439 7.611723 7.624282 CJM27 7.804139 7.578639 7.695482 3 CJM29 7.0086 7.424882 6.968483 7.468347 CJM30 7.935507 7.606381 7.576341 7.655138

[0040] Table 3

[0041]

[0042]

[0043] 2. Rescreening

[0044] According to the initial screening results, four new yeast strains were selected, and SC16-1 ammonium sulfate medium was used as a control. Liquid fermentation was performed, and the relevant indicators were tested at 12h, 24h, 36h, 48h, 60h, and 72h from 0h. The strains, culture medium, and operation are shown in Table 4.

[0045] Table 4

[0046]

[0047] FJM14 made good use of ammonia water and ammonium sulfate; FJM18 also made good use of ammonia water and ammonium sulfate; the amount of protein produced in 1L culture medium was 1.5g / L; the utilization rate of nitrogen in ammonia water by FJM18 was 9.39%.

[0048] Detection of nitrogen content in fermentation sludge and centrifugal supernatant:

[0049] The calculation method of bacterial protein is as follows: the bacterial sludge obtained by centrifuging all the fermentation liquid in each bottle of the sample is used to determine the nitrogen content (g) in the bacterial sludge using the Kjeldahl method.

[0050] Nitrogen content in bacterial sludge (g) × 6.25 = protein content in bacterial protein (g);

[0051] Protein content in bacterial protein (g) × 1000 / sample per bottle of fermentation broth (mL) = protein content of fermentation broth (mg / mL, i.e. g / L);

[0052] The conversion rate of non-protein nitrogen is calculated as follows: nitrogen content of all bacterial sludge in each bottle of sample (g) / [amount of non-protein nitrogen added to each bottle of fermentation liquid of sample (g) × nitrogen content of non-protein nitrogen (%)] × 100 = conversion rate of non-protein nitrogen (%).

[0053] The results of the protein content g / L (bacterial mud) in the fermentation broth are shown in Table 5, and the results of the nitrogen content g / L (bacterial mud) in the centrifugal supernatant are shown in Table 6.

[0054] Table 5

[0055] Time 0 12 24 36 48 60 72 SC16-1 Ammonium sulfate 0 0.036 0.044 0 0 0.006 0 FJM14 Ammonium sulfate 0 0.195 0.604 0.883 1.091 1.158 1.259 Ammonium chloride 0 0.050 0.358 0.510 0.647 0.769 0.891 Urea 0 0.086 0.211 0.268 0.303 0.351 0.348 ammonia 0 0.048 0.510 0.920 1.135 1.313 1.413 FJM15 Ammonium sulfate 0 0 0 0 0 Ammonium chloride 0 0 0 0 Urea 0 ammonia 0 0 0 Ammonium bicarbonate 0 0 FJM16 Ammonium sulfate 0 0.035 0.025 0.268 0.679 0.809 0.788 Ammonium chloride 0 0.033 0.028 0.211 0.362 0.393 0.413 Urea 0 0.047 0.147 0.374 0.414 0.438 0.442 ammonia 0 0.036 0.030 0.486 0.876 0.952 0.982 Ammonium bicarbonate 0 0.071 0.082 FJM18 Ammonium sulfate 0 0.213 0.655 0.876 0.978 1.077 1.253 Ammonium chloride 0 0.138 0.454 0.639 0.783 0.993 1.145 Urea 0 0.153 0.223 0.278 0.329 0.356 0.399 ammonia 0 0.185 0.633 0.946 1.133 1.266 1.522

[0056] Table 6

[0057]

[0058]

[0059] From Tables 5 and 6, it can be seen that FJM60 Candida utilis and FJM18 Pichia kudriavzevii have the strongest ability to utilize non-protein nitrogen, with the number of viable bacteria reaching 10 7 cfu / mL. The changes of sample bacterial protein content, wet weight and viable count were basically consistent.

[0060] The growth curves of FJM18 and FJM60 in the basal and ammonia water medium were measured every 2 hours for 30 hours. The experiment was repeated twice (three times in each group), and the results were basically consistent with those of every 2 hours and every 4 hours. The subsequent culture time was selected from the logarithmic phase of 15-18 hours. The growth curves of FJM18 and FJM60 in the basal ammonia water medium are shown in Figure 2. Figure 1-2 As shown, Figure 1 Sampling is done every 2 hours. Figure 2 Sampling was performed every 4 hours.

[0061] Example 2 Single yeast fermentation experiment

[0062] 1. Single yeast fermentation experiment of distiller's grains

[0063] FJM18 fermented three kinds of Luzhou and Wuliangye (and dried part of the wet lees and crushed them) wet material fermentation, and calculated the inoculation amount of yeast solution (ammonia culture medium) with an initial OD value of 0.5. The amount of ammonia added was 100g + 2.75ml ammonia, and the pH was 6.8-7. The inoculation amount and the number of live bacteria in the culture solution are shown in Table 7. The wet material was sampled once every 24 hours (20 g) and dried. The results of the number of live yeast corresponding to the solid-state fermentation time of the single bacteria of Luzhou Laojiao Luzhou-flavor lees are shown in Table 8. The results of the number of live yeast corresponding to the solid-state fermentation time of the single bacteria of Wuliangye Luzhou-flavor lees are shown in Table 9.

[0064] Table 7

[0065]

[0066] Table 8 Table 9

[0067]

[0068] From the data in Tables 8 and 9, it can be seen that when the FJM18 strain was fermented with single yeast on two different types of lees, Luzhou Laojiao and Wuliangye, the trend of the change of the number of live bacteria over time was different. On the Luzhou Laojiao lees, the number of live bacteria of the FJM18 strain increased from the initial 1.6×10 6 cfu / mL increased to 1.76×10 at 120 hours 8 cfu / mL, showing a clear growth trend. On Wuliangye distiller’s grains, the number of viable bacteria decreased to 1.3×10 6 cfu / mL, and then rapidly increased to 3.3×10 at 120 hours. 8 This may indicate that the environment of Wuliangye distiller's grains had a certain inhibitory effect on the initial growth of FJM18, but then the strain adapted to the environment and began to multiply in large quantities.

[0069] The data of the control group of natural fermentation showed that the number of microorganisms in the lees was low without inoculation of FJM18 strains, and contamination by foreign bacteria occurred during the fermentation process, especially in Wuliangye lees, where the number of foreign bacteria reached 3.3×10 8 cfu / mL, which may have a negative impact on the fermentation quality of the lees.

[0070] In summary, the FJM18 strain can effectively grow on two different types of distiller's grains, but its growth rate and adaptability to the environment vary in different distiller's grains. In Luzhou Laojiao distiller's grains, the growth of the FJM18 strain is more stable, while in Wuliangye distiller's grains, although the initial growth is slow, the later growth is rapid, showing a strong adaptability. The control group data of natural fermentation also suggests the importance of inoculating specific strains in actual production to ensure the stability of the fermentation process and the quality of the product.

[0071] The crude protein of the liquor lees fermented by FJM18 was tested. The results of the Luzhou Laojiao Luzhou-flavor fermentation are shown in Table 10, and the results of the Wuliangye Luzhou-flavor fermentation are shown in Table 11.

[0072] Table 10

[0073]

[0074] Table 11

[0075]

[0076] In Tables 10 and 11, we can see the fermentation effect of FJM18 strains in different Luzhou-flavor distiller's grains. In Luzhou Laojiao Luzhou-flavor distiller's grains, the crude protein content of FJM18 strains increased during the 48-hour and 96-hour fermentation process, indicating that FJM18 strains can effectively utilize the nutrients in the distiller's grains for growth and metabolism. However, during the natural fermentation process, due to contamination by foreign bacteria, the number of live bacteria decreased significantly after 96 hours, and the crude protein content also decreased, indicating that foreign bacteria contamination had a negative impact on the fermentation process.

[0077] In Wuliangye Luzhou-flavor distiller's grains, FJM18 strains also showed good fermentation effects during the 48-hour and 96-hour fermentation processes, and the crude protein content increased. Compared with Luzhou Laojiao Luzhou-flavor distiller's grains, the fermentation effect of FJM18 strains in Wuliangye Luzhou-flavor distiller's grains was more significant, especially in terms of crude protein content, which reached 14.173% after 96 hours, higher than the 13.111% of Luzhou Laojiao Luzhou-flavor distiller's grains. This may be related to the raw material characteristics of Wuliangye Luzhou-flavor distiller's grains, which may be more suitable for the growth and metabolism of FJM18 strains.

[0078] Combining the data in the two tables, we can conclude that the fermentation effect of FJM18 strain in different Luzhou-flavor distiller's grains varies, but overall it can effectively improve the nutritional value of distiller's grains. However, in order to ensure the stability of the fermentation process and the quality of the product, the fermentation environment must be strictly controlled to prevent contamination by foreign bacteria. Future research can further explore how to optimize the fermentation conditions to improve the application effect of FJM18 strain in distiller's grains fermentation.

[0079] 2. Single yeast fermentation experiment of wheat bran

[0080] 3% urea and 3% ammonium sulfate were added as nitrogen sources to the wheat bran treated by the FJM18 fermentation process. Under the condition of an initial OD value of 0.5, an appropriate amount of yeast liquid was added and fermented for 48 hours. Subsequently, the wet material after fermentation was dried. The specific contents of dry matter recovery, crude protein, true protein and other nutrients of the fermentation group with 3% urea and 3% ammonium sulfate are shown in Table 12.

[0081] Table 12

[0082]

[0083]

[0084] Wheat bran was used as the fermentation raw material, and 1%, 2% and 3% of ammonia water, urea and ammonium sulfate were added thereto respectively. Yeast liquid was added under the condition of initial OD value of 0.5, and fermentation treatment was carried out for 48 hours. Subsequently, the wet material after fermentation was dried, and the content of crude protein and true protein was tested. The experimental results are shown in Table 13.

[0085] Table 13

[0086]

[0087] Single-bacteria fermentation experiments were carried out using FJM18 using wheat bran and distiller's grains as substrates through solid-state fermentation. A comprehensive evaluation was conducted on the true protein, NPN utilization efficiency, and mannan content in the bacterial protein products of the single bacteria.

[0088] First, using wheat bran as a fermentation substrate, after solid-state fermentation treatment by FJM18, it was found that its true protein content was significantly improved, and the NPN utilization efficiency was also improved. This shows that FJM18 can effectively utilize the nitrogen source in the substrate during wheat bran fermentation to promote the synthesis of bacterial protein. In addition, the present invention also found that the mannan content in the fermented wheat bran was also increased, which further confirmed the positive role of FJM18 in solid-state fermentation.

[0089] Next, solid-state fermentation was performed using FJM18 with white wine lees as the fermentation substrate. The experimental results showed that the true protein content and NPN utilization efficiency in white wine lees were also significantly improved. This indicates that FJM18 also has a good application effect in the fermentation process of white wine lees.

[0090] In summary, single-bacteria fermentation using FJM18 by solid-state fermentation can effectively increase the true protein content in the substrate and the utilization efficiency of NPN.

[0091] Example 3 Evaluation of the nutritional value of fermented feed

[0092] The key nutritional indicators of the fermented wheat bran in Example 2 were measured.

[0093] 1. Index determination method

[0094] Dry matter: Refer to GB / T 6435-2014 Determination of moisture in feed and use a constant temperature air drying oven for determination.

[0095] Crude protein: Refer to GB / T 6432-2018 crude protein determination method in feed, and use FOSS fully automatic Kjeldahl nitrogen analyzer for determination.

[0096] True protein: Determined by copper sulfate method using FOSS fully automatic Kjeldahl nitrogen analyzer.

[0097] Crude fiber: refer to DB37 / T 3370-2018 method for determination of crude fiber content in feed, and use polyester mesh bag-automatic fiber analyzer for determination.

[0098] Energy: measured using an IKA C6000 oxygen bomb calorimeter.

[0099] Reducing sugar: DNS method was used.

[0100] Organic acids: using high performance liquid chromatography.

[0101] 2. Determination of apparent metabolizable energy value of fermented wheat bran in poultry

[0102] Test method: Select Hy-Line brown shell roosters weighing (2.5±0.2) kg to conduct gavage emptying tests on various feed raw materials according to GB / T26437-2010 "Technical regulations for the determination of apparent metabolizable energy of chicken feed by gavage for evaluation of the effectiveness and safety of livestock and poultry feeds". Each raw material was repeated 8 times, and each repeat had 1 chicken. At least one week before the start of the metabolic test, each chicken underwent excrement collection bottle suture surgery and recovered to normal body condition. Each raw material was tested in three stages: pre-feeding period (3d), test period (48h fasting emptying, gavage, 48h collection of excrement) and body condition recovery period. The start time of fasting emptying was accurately recorded, and water was freely available during the fasting period. After the fasting ended, each chicken was accurately gavaged with 25-mesh air-dried feed (preparation method: fermented wheat bran was directly dried in the open air, and the wet material was evenly air-dried) 50g through a gavage, and the completion time of gavage was recorded by individual. All excrement was collected 48 hours after force-feeding, dried at 65°C and then rehydrated for 24 hours. The gross energy and amino acid content were determined and the apparent metabolizable energy of the feed raw materials was calculated.

[0103] The apparent metabolizable energy (AME, in MJ / kg) of feed for each replicate group of chickens was calculated as follows:

[0104]

[0105] GE1=E1×M1;

[0106] GE2=E2×M2.

[0107] Where:

[0108] GE1——total energy intake, in joules (J);

[0109] GE2——gross excrement energy, in joules (J);

[0110] E1——energy value of dry matter intake, in joules per gram (J / g);

[0111] M1——dry matter intake, in grams (g);

[0112] E2——energy value of dry matter of excrement, in joules per gram (J / g);

[0113] M2——Dry matter mass of excrement 48 hours after ingestion of the tested feed, in grams (g). The results of key indicators of fermented wheat bran (air-dried basis) are shown in Table 14.

[0114] Table 14

[0115]

[0116]

[0117] The apparent metabolizable energy value of fermented wheat bran for laying roosters was (10.74±2.98) MJ / kg, which was 25.9% higher than that of unfermented wheat bran (8.53±0.03) MJ / kg. In addition, after fermentation, the content of true protein, reducing sugar and organic acid in wheat bran was significantly increased. This result fully proves the positive role of FJM18 in improving the nutritional value of wheat bran.

[0118] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention rather than to limit it. Although the present invention has been described in detail with reference to the above embodiments, ordinary technicians in the relevant field should understand that the specific implementation methods of the present invention can still be modified or replaced by equivalents. Any modification or equivalent replacement that does not depart from the spirit and scope of the present invention should be covered within the scope of protection of the claims of the present invention.

Claims

1. A Pichia kudriavzevii FJM18, characterized in that The deposit number of the Pichia kudriavzevii FJM18 is CGMCC No.30405.

2. An application of the Pichia kudriavzevii FJM18 according to claim 1, characterized in that: The application includes at least one of the following: A) application of the Pichia kudriavzevii FJM18 in fermenting wheat bran; B) application of the Pichia kudriavzevii FJM18 in fermentation of distiller's grains; C) Application of the Pichia kudriavzevii FJM18 in the preparation of biological feed additives; D) Application of the Pichia kudriavzevii FJM18 in preparing biological feed; E) Application of the Pichia kudriavzevii FJM18 in degrading organic pollutants; F) Application of the Pichia kudriavzevii FJM18 in improving the apparent metabolizable energy of poultry.

3. The use according to claim 2, characterized in that: The organic pollutant is wheat bran or distiller's grains.

4. A bacterial agent, characterized in that The bacterial agent includes the Pichia kudriavzevii FJM18 described in claim 1.

5. A method for preparing bacterial protein feed, characterized in that: include: The fermentation substrate is fermented using the Pichia kudriavzevii FJM18 described in claim 1 to obtain the bacterial protein feed.

6. The method for preparing bacterial protein feed according to claim 5, characterized in that: The fermentation substrates are wine lees and ammonia water.

7. The method for preparing bacterial protein feed according to claim 5, characterized in that: The fermentation substrates are wheat bran, urea and ammonium sulfate.

Images