Search Results (18)

Consumer interest in vegetarian, ethical, and clean-label foods is reviving the use of plant-derived milk coagulants. Cardosins from Cynara cardunculus (“thistle”) are aspartic proteases with strong clotting activity, yet their technological impact in cheese remains under-explored. This study compared a commercial thistle extract (PC) with traditional bovine rennet rich in chymosin (AC) during manufacture and 60-day ripening of Caciotta cheese. Classical compositional assays (ripening index, texture profile, color, solubility) were integrated with scanning electron microscopy, three-dimensional surface reconstruction, and descriptive sensory analysis. AC cheeses displayed slower but sustained proteolysis, yielding a higher and more linear ripening index, softer body, greater solubility, and brighter, more yellow appearance. Imaging revealed a continuous protein matrix with uniformly distributed, larger pores, consistent with a dairy-like sensory profile dominated by milky and umami notes. Conversely, PC cheeses underwent rapid early proteolysis that plateaued, producing firmer, chewier curds with lower solubility and darker color. Micrographs showed a fragmented matrix with smaller, heterogeneous pores; sensory evaluation highlighted vegetal, bitter, and astringent attributes. The data demonstrate that thistle coagulant can successfully replace animal rennet but generates cheeses with distinct structural and sensory fingerprints. The optimization of process parameters is therefore required when targeting specific product styles. Full article

Plant-derived rennet sources have been recognized as effective alternatives to traditional animal-derived rennet in cheese processing, producing various cheeses for vegetarians and Halal consumers. The aim of the present study was to assess the quality and identify potential chemical and sensory markers of Canestrato cheese made from pasteurized mixed cow/sheep milk (50:50 ratio) using a kiwifruit extract (prepared in the laboratory) as a rennet source of vegetable origin. The milk was curdled with an aqueous kiwi extract (cK group), with commercial calf rennet (cT group), or with commercial microbial rennet obtained from the controlled fermentation of Rhizomucor miehei (cH group). For each cheesemaking process, 30 L of mixed milk was used, resulting in 10 forms, each weighing approximately 1 kg. Cheese produced using kiwifruit rennet exhibited improved nutritional characteristics, such as lower total lipid, cholesterol, and sodium content and greater levels of polyphenols, potassium, and phytosterols compared with the cheese of the cT group. The cheese of the cK group had a slightly more pronounced bitterness compared to cheeses made with conventional rennets (cH and cT). Overall, the use of kiwifruit extract as a milk coagulant has proved to be a promising alternative, improving both the nutritional profile and functional properties of the resulting cheese. Full article

This study explores the extraction and characterization of proteolytic enzymes from brewer’s spent grain (BSG) and their potential as sustainable coagulants in the dairy industry. BSG samples from various beer types (Blonde Ale, IPA, Kölsch, Honey, and Porter) were obtained from two artisanal breweries in Mar del Plata, Argentina. Optimization of caseinolytic activity (CA) and protein extraction was conducted using a Plackett–Burman design, followed by a Box–Behnken design. Optimal protein concentration was achieved at intermediate pH and high temperature, while CA peaked at pH 8.0. The specific caseinolytic activity (SCA) varied among the extracts, with BSG3 showing the highest activity (99.6 U mg⁻¹) and BSG1 the lowest (60.4 U mg⁻¹). Protease inhibitor assays suggested the presence of aspartic, serine, metallo, and cysteine proteases. BSG3 and BSG4 showed the highest hydrolysis rates for α-casein (70% and 78%). For κ-casein, BSG1, BSG2, and BSG3 demonstrated moderate activity (56.5%, 49%, and 55.8), while BSG4 and BSG5 exhibited the lowest activity. Additionally, the milk-clotting activity (MCA) of BSG extracts was comparable to plant-based coagulants like Cynara cardunculus and Ficus carica. These findings highlight the potential of BSG-derived proteases as alternative coagulants for cheese production, offering a sustainable link between the brewing and dairy industries. Full article

The microbiological and biochemical properties of a goat cheese produced using Helianthus annuus (sunflower) seed extract as a coagulant and the potentially probiotic autochthonous culture Limosilactobacillus mucosae CNPC007 were examined in comparison to a control cheese devoid of the autochthonous culture. Throughout a 60-day storage period at 6 ± 1 °C, lactobacilli maintained a count of above 8 log CFU/g. Additionally, its viability in cheeses subjected to the in vitro gastrointestinal conditions demonstrated improvement over this period. Specifically, the recovery of lactobacilli above 6 log CFU/g was observed in 16.66% of the samples in the first day, increasing to 66.66% at both 30 and 60 days. While total coliforms were detected in both cheese trials, this sanitary parameter exhibited a decline in L. mucosae cheeses during storage, falling below the method threshold (<3 MPN/g) at 60 days. This observation suggests a potential biopreservative effect exerted by this microorganism, likely attributed to the higher acidity of L. mucosae cheeses at that point (1.80 g/100 g), which was twice that of the control trial (0.97 g/100 g). Furthermore, distinct relative proportions of >30 kDa, 30–20 kDa, and <20 kDa proteins during storage was verified for L. mucosae and control cheeses. Consequently, either the H. annuus seed extract or the L. mucosae CNPC007 autochthonous culture influenced the biochemical properties of the cheese, particularly in terms of proteolysis. Moreover, L. mucosae CNPC007 acidification property resulted in a biopreservative effect throughout the storage period, indicating the potential as a promising source of probiotics for this product. Full article

Onopordum nervosum subsp. platylepis Murb. is an Asteraceae species found in Tunisia, Algeria, and Libya. It has been studied for its potential use as a vegetable rennet alternative to animal-derived rennet, making it important to understand its germination characteristics. This species often shows low germinability due to dormancy, which limits its large-scale cultivation. In preliminary experiments, many factors were analyzed to evaluate the seed germination of this thistle including physical stratification with abrasive paper and high-temperature water, chemical treatments with hydrogen peroxide (H₂O₂) or sulfuric acid (H₂SO₄), and the exogenous use of the growth regulator gibberellic acid (GA₃). Based on the obtained results and subsequent analysis, GA₃ and cold stratification durations were selected for the second experiment, which used a full factorial RSM design with three levels for each factor. Additionally, the total polyphenol content (TPC), total flavonoid content (TFC), condensed tannin content (CTC), and DPPH assay were analyzed during the optimization process and for ungerminated seeds. The findings revealed that 14 days of cold stratification followed by 750 ppm GA₃ was the most efficient method for breaking the dormancy of these thistle seeds. The highest TPC was recorded in ungerminated seeds. However, germinated Onopordum nervosum ssp. platylepis seeds showed higher levels of TFC and CTC (14.83 mg QE/g DW and 6.49 mg EC/g DW, respectively) compared to the non-germinated ones and demonstrated the greatest potency in inhibiting free radicals (DPPH EC₅₀ = 0.018 mg/mL) at the identified optimal germination conditions. Ungerminated seeds indicated significant phenolic content (TPC) and a limited ability to reduce oxidants, which could explain their low germination percentage. Our findings on the seed germination and dormancy characteristics of this endemic thistle will aid in the protection and development of its germplasm. Full article

This study aimed to compare the chemical composition of aqueous extracts from different aerial plant parts (leaves, stems, and flowers) of Onopordum nervosum ssp. platylepis growing in different regions (Sousse, Kairouan, and Nabeul) in Tunisia, as well as their antioxidant and milk-clotting properties for both spontaneously grown and cultivated plants. Results showed that phenolic composition varies significantly among ecotypes and plant organs (p < 0.05), with flowers containing the highest amounts of total phenols and flavonoids in both plant types. The flowers from Nabeul (NA) region showed the highest amounts of total phenols and higher phenolic contents compared to leaves and stems, with 44.75 mg GAE/g and 39.79 mg GAE/g in spontaneous and cultivated plants, respectively. However, flowers of spontaneously plants grown in Sousse (SO) showed the highest total flavonoid contents (11.42 mg QE/g). Additionally, the findings indicated that flowers contained higher concentrations of mono- and disaccharides than leaves and stems. The antioxidant activity showed that the radical scavenging activity of O. platylepis aqueous extracts is significantly affected by the organ and genotype (p < 0.05). NA genotype revealed the highest potency in inhibiting free radicals, with flowers having the lowest IC50s values in 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) tests, registering 0.13 ± 0.02 and 0.14 ± 0.01 mg/mL, respectively. Milk-clotting activity was limited to flowers of this thistle. Moreover, SO and NA genotypes were identified as the most potent populations to coagulate milk in spontaneous and cultivated plants. The principal component analysis confirms the organ and genotype variability in this thistle, with the first two axes explaining 64.15% of the variance and highlighting a distinct flower group. The obtained results suggest that the domestication of this thistle could be useful for the conservation of biodiversity and the promotion of genotypes with potential technological properties. Full article

Agroindustrial by-products constitute an alternative source of feed livestock, and their use contributes to the sustainability of livestock systems and the circular bioeconomy. The effects of replacing cereal (0%, 40%, and 80%) with dehydrated orange pulp (DOP) in the diet of goats on the antioxidant and fatty acid (FA) contents of cheeses were evaluated. For a more suitable understanding of the role of coagulant enzymes in establishing the properties of the cheese, the effect of milk-clotting with animal and vegetable rennet was also analysed. The rennet did not substantially affect the FA or the antioxidant compounds, and the use of DOP did not affect the FA contents. However, the α-tocopherol levels, total phenolic compounds (TPC), and total antioxidant capacity (TAC) in cheeses increased as the percentage of DOP replacing cereals increased. Moreover, the high correlation obtained between the TAC and the TPC (r = 0.73) and α-tocopherol (r = 0.62) contents indicated the important role played by these compounds in improving the antioxidant capacity of the cheese. In conclusion, DOP is a suitable alternative to cereals in the diet of goats and improves the antioxidant status of the cheese produced. Full article

Cheese production is an ancient practice to preserve a perishable food, such as milk, for a long time. The first step of cheese processing involves the addition of rennet, which contains the enzymes necessary for the hydrolysis and coagulation of the caseins present in milk. Typically, animal-derived rennet, such as calf rennet containing chymosin, are used as source of enzymes for cheese processing. Alternatively, microbial chymosin or recombinant chymosin is used. However, recently, plant-derived rennet such as the ones derived from thistle and bitter orange flowers and from artichoke (Cynara cardunculus var. scolymus) have also been demonstrated to be valid sources of enzymes for cheese processing. Therefore, herein, different plant and fruit extracts were tested and compared for their ability to coagulate milk caseins. In particular, beyond artichoke and cardoon (Cynara cardunculus) extracts, those from pineapple (Ananas comosus (L.) Merr.), papaya (Carica papaya L.), common fig (Ficus carica L.) milky sap, and oyster mushroom (Pleurotus ostreatus (Jacq. ex Fr.) P. Kumm.) were investigated for their proteolytic, esterase, and milk-clotting activities. The extracts were then exploited as vegetable and fruit rennet for the experimental production of cheeses, which were examined, after 30 days of maturation, for their moisture, fat, protein, and free fatty acid (FFA) content. Interestingly, the artichoke, cardoon, and thistle mushroom extracts showed high proteolytic activity compared to calf rennet, while the level of esterase activity appeared to be similar for all the extracts. The papaya extract showed the lowest proteolytic and esterase activity. Although the pH, moisture, fat, and protein contents were very similar to those of cheese made with calf rennet, the medium- and long-chain FFAs broadly differed among produced cheeses, with variations in the lipid quality indices. Full article

Onopordum tauricum Willd., a species distributed in Eastern Europe, has been the subject of various research endeavors aimed at assessing its suitability for extracting vegetable rennet for use in the production of local cheeses as a substitute for animal-derived rennet. In Italy, the species has an extremely fragmented and localized distribution in six locations scattered across the central-northern Apennines and some areas of southern Italy. In this study, both the morphology and genetic diversity of the six known Italian populations were investigated to detect putative ecotypes. To this end, 33 morphological traits were considered for morphometric measurements, while genetic analysis was conducted on the entire genome using the ddRAD-Seq method. Both analyses revealed significant differences among the Apennine populations (SOL, COL, and VIS) and those from southern Italy (ROT, PES, and LEC). Specifically, the southern Italian populations appear to deviate significantly in some characteristics from the typical form of the species. Therefore, its attribution to O. tauricum is currently uncertain, and further genetic and morphological analyses are underway to ascertain its systematic placement within the genus Onopordum. Full article

Milk coagulation is a process used for the formulation of different dairy products such as cheese. In this process, milk undergoes changes in its chemical stability thanks to acidification or enzymatic reactions. Traditionally, milk coagulation has been carried out with rennet of animal origin, but recently, the research of new types of rennet such as microbial rennet and vegetable rennet has increased. This study aims to present an organized review of the most relevant information on lactic coagulation, its relationship with vegetable rennets, and the importance of the botanical genus Cynara in the extraction of vegetable rennets, focusing on the coagulant potential of artichoke (Cynara scolymus). We conducted this literature review and found that lactic coagulation and vegetable rennets are linked through the enzymatic activity of the latter. The results of the main studies demonstrated a strong relationship between vegetable rennets and protease enzymes as well as the presence of these enzymes in extracts of cardoon (Cynara scolymus) and artichoke (Cynara scolymus). In addition, studies highlight the presence of thistle extracts in artisanal cheese preparations in the Iberian Peninsula. Based on the results of the studies, a comparison between cheeses made with vegetable rennet and those made with traditional rennet was also carried out. Although the results show that the use of vegetable rennet in the manufacture of cheese can confer undesirable characteristics, the use of extracts from Cynara plants demonstrates that vegetable rennets have an industrial potential, especially the one obtained from artichoke (Cynara scolymus) due to its high availability. Nevertheless, specific studies are required for a better understanding and application of this rennet. Full article

Milk coagulants prepared by maceration of flowers harvested from both spontaneous and cultivated Onopordum tauricum Willd. and a commercially available coagulant obtained from Cynara cardunculus L. (control) were assayed for small-scale manufacturing of Caciofiore, an Italian specialty raw ewe’s milk cheese produced in a family run dairy farm located in the Marche region (Central Italy). The microbiota of the three thistle-based milk coagulants and their effect on the microbial dynamics of raw milk cheeses during fermentation and maturation (from day 0 up until day 60) were investigated through a combined approach based on viable counting and Illumina DNA sequencing. In both the control and experimental cheeses, despite the slight differences emerged depending on the coagulant used, Lactococcus lactis and Debaryomyces hansenii were the prevalent species among bacteria and fungi, respectively. Moreover, raw ewe’s milk was the main factor affecting the evolution of both the bacterial and fungal microbiota in all cheeses. The overall similarities between control and experimental cheeses herein analyzed supports the exploitation of Onopordum tauricum Willd. as an alternative milk coagulating agent for production of Caciofiore and, more in general, raw ewe’s milk cheeses. Full article

The aim of this study was to identify novel milk coagulants to be used in cheesemaking. For this purpose, aqueous extracts from safflower (Carthamus tinctorius), sunflower (Helianthus annuus), flax (Linum usitatissimum) and sesame (Sesamum indicum) seeds were tested for their caseinolytic (CA) and milk coagulating properties (MCA) in skim milk at temperatures of 25, 37, 50, 65 and 80 °C. The seed oil samples with the highest temperature ranges in regard to coagulation efficiency were then tested in cow, buffalo, goat and sheep milks and the MCA and curd yield (CY) parameters were measured at different temperatures. Due to their high milk coagulation efficiency (CE) in all types of milk and at different temperatures, the sesame and sunflower seed extracts proved to be particularly interesting and their CY parameters were similar to those obtained with animal rennet. Moreover, our results confirm that oilseed coagulants are capable of coagulating milk and can also be considered as potential animal rennet substitutes. This study provides valuable insights into the development of potential vegetable coagulants that could be used for various production processes aimed at specific target consumers. Full article

The reduced availability and the increasing prices of calf rennet, coupled to the growing global demand of cheese has led, worldwide, to explore alternative clotting enzymes, capable to replace traditional rennet, during the cheesemaking. In addition, religious factors and others related to the vegetarianism of some consumers, have led to alternative rennet substitutes. Nowadays, several plant-derived milk-clotting enzymes are available for cheesemaking technology. Many efforts have also been made to compare their effects on rheological and sensory properties of cheese to those arising from animal rennet. However, vegetable clotting enzymes are still partially suitable for cheesemaking, due to excessive proteolytic activity, which contribute to bitter flavor development. This review provides a literature overview of the most used vegetable clotting enzymes in cheese technology, classified according to their protease class. Finally, clotting and proteolytic activities are discussed in relation to their application on the different cheesemaking products. Full article

Australian sweet lupin, the largest legume crop grown in Western Australia, is receiving global attention from the producers of new foods. To understand the effect of protein on cheese yield, lupin milk proteins were separated from the first, second, and third filtrations by cheesecloths. However, proteins from the first and second were analyzed using two-dimensional polyacrylamide gel electrophoresis; then, the isolated proteins associated with cheese production were identified. The research also focused on identifying the optimal method of cheese production based on the coagulation process, temperature, yield, and sensory evaluation. Lupin curds from the two cultivars, Mandelup and PBA Jurien, were produced using vinegar, lemon juice, starter culture, vegetable rennet enzyme as coagulant, as well as curd generated using starter culture and vegetable rennet enzyme. Cow’s milk was used as a control. The results indicated that first-time filtration produced better extraction and higher yield of lupin proteins and cheese than the second filtration. A sensory analysis indicated that lupin cheese produced from PBA Jurien lupin milk using vinegar, 7.80% expressed as acetic acid, and ground in 45 °C water, was the most acceptable. The cheeses were examined for their protein, carbohydrates, fat, ash, and moisture contents. The concentration of protein was approximately 27.3% and 20.6%, respectively, in the cheese from PBA Jurien and Mandelup. These results suggest that lupin milk can adequately supply the proteins needed in human diets and, thus, could be used in the production of many existing products that require animal milk as an input. Full article

In Mediterranean cropping systems, it is important to utilize marginal lands for the cultivation of non-food crops. Spontaneous thistle genera such as Cynara cardunculus L. and Onopordum tauricum Willd. are native to southern Europe. Previous research has focused on their spontaneous growth in the environment or ability to supply biofuel. The aim of this study was to identify the effect of two different planting densities on the flower differentiation, vegetable rennet production and weed control of O. tauricum Willd. and C. cardunculus L. in rainfed unfertilized Mediterranean cropping systems. The results showed that plant density influenced most of the biomass parameters for O. tauricum Willd. but no significant differences were observed for C. cardunculus L. Higher densities of both species were beneficial for weed control. We recommend the use of C. cardunculus L. and O. tauricum Willd. as suitable species for vegetable rennet production in Mediterranean organic cropping systems. Full article