Trehalose: Physicochemical Characteristics, Manufacturing Processes, and Application Guidelines
Trehalose: Physicochemical Characteristics, Manufacturing Processes, and Application Guidelines
Trehalose is a non-reducing disaccharide composed of two glucose units linked by an α,α-1,1-glycosidic bond. It exhibits good chemical stability and formulation compatibility across a range of processing and storage conditions. Owing to its moisturizing capability, anti-freezing and desiccation tolerance, and stabilizing effects on proteins and membrane systems, trehalose has attracted broad attention in foods, pharmaceuticals and bioproducts, cosmetics, and agriculture.
Keywords: Trehalose; Non-reducing disaccharide; Vitrification; Lyoprotectant; Enzymatic synthesis; Microbial fermentation; Stabilizer
I. Research Brief History
1.1 Discovery and Industrialization
In 1832, the British scientist Wiggers first isolated trehalose from ergot fungus. Subsequently, trehalose production evolved from extraction of natural sources to recovery from microbial systems and fermentation, as well as enzymatic conversion routes using starch as the substrate, enabling scalable manufacturing. China achieved industrial-scale trehalose production starting in 2000.
1.2 Form Types and Application Relevance
Based on the presence or absence of crystal water, trehalose can be categorized as anhydrous trehalose or crystalline trehalose. Different forms may differ in moisture control, powder flowability, hygroscopic behavior, and stability in certain formulations. Selection of the material form should align with the target process (e.g., spray drying, freeze-drying, or powder blending) and storage conditions.
1.3 Sweetness and Processing Stability
Trehalose generally has a lower sweetness than sucrose and provides a mild sweetness profile, while maintaining good thermal and acid stability. As a non-reducing sugar, trehalose is less likely to participate in typical browning reactions (Maillard reactions) in the presence of amino acids or proteins, even under heating, which is advantageous for systems where color and flavor drift must be controlled.
II. Basic Information and Physicochemical Parameters
2.1 Key Information Summary
Item | Information | Item | Information |
Name | D-Trehalose anhydrous | Density | 1.512 g/cm³ |
Molecular Formula | C12H22O11 | Appearance | White to off-white powder |
Molecular Weight | 342.297 | Flash Point | 362.3 °C |
CAS No. | 99-20-7 | Applications | Food processing; pharmaceuticals; cosmetics; agriculture |
EINECS No. | 202-739-6 | Safety Description | Stable properties |
Melting Point | 203 °C | Hazard Symbol | Xi |
Boiling Point | 675.4 °C | Hazard Statement | R38 |
Water Solubility | 68.9 g/100 g water (25 °C) | RTECS No. | LZ5776770 |
|
| HS Code | 2940009000 |
2.2 Structural Features and the Significance of Non-reducing Behavior
The α,α-1,1-glycosidic bond in trehalose confers a non-reducing character. In formulations containing proteins or amino compounds, this property can reduce the likelihood of participation in typical browning pathways during thermal processing, and may help mitigate unintended reactions involving protein amino groups, thereby supporting color and flavor stability management.
III. Properties and Technical Value Foundations
3.1 Stability and Safety
(1) Non-reducing Character and Browning Risk Control
Trehalose is non-reducing and, in systems containing amino acids or proteins, typically does not readily trigger classic Maillard browning pathways, which helps maintain color and flavor consistency during heating or storage. It should be noted that color and flavor changes in real formulations may also arise from lipid oxidation, aroma volatilization, or protein structural changes, and therefore should be evaluated in the context of the specific process and formulation conditions.
(2) Metabolism and Tolerance Boundaries
After entering the gastrointestinal tract, trehalose can be hydrolyzed by trehalase into glucose and then enters conventional carbohydrate metabolic pathways. Statements regarding safety and health effects should be bounded by the permitted regulatory context, use levels, and actual exposure. At higher intakes or in special populations, inter-individual differences in digestive enzyme activity should also be considered when evaluating tolerance.
3.2 Low Hygroscopicity
Trehalose typically exhibits low hygroscopicity, and its moisture uptake behavior is influenced by crystal form, water content, and ambient relative humidity. In food and pharmaceutical powder systems, this property can help reduce risks of moisture-induced caking, loss of flowability, and physical drift caused by moisture regain. For high-humidity storage or moisture-sensitive systems, it is recommended to incorporate crystal form selection and moisture specifications into raw material release criteria and in-process controls.
3.3 High Glass Transition Temperature and Process Implications
Compared with certain other disaccharides, trehalose has a relatively high glass transition temperature. Under low-moisture conditions, it is therefore more prone to form a glassy state, reducing molecular mobility in the solid phase. This behavior, together with its process stability and low hygroscopicity, contributes to its value in flavor retention during spray drying and in stabilization of high-protein systems. Glass formation and stabilization performance are highly dependent on moisture content, co-solute composition, and processing temperature history, and should be confirmed via thermal analysis and accelerated stability evaluation.
3.4 Non-specific Protective Effects on Biomacromolecules and Biological Systems
(1) Protein Systems
Trehalose can reduce tendencies for protein denaturation and aggregation under freezing or drying stress by modulating the hydrogen-bond network of the hydration layer and by forming a glassy matrix in the solid state, thereby supporting conformational stability.
(2) Membrane and Lipid Systems
Trehalose can form hydrogen-bond interactions with phospholipid headgroups, helping maintain membrane integrity under dehydration or freezing conditions and reducing risks of membrane fusion and abnormal permeability. These effects are sensitive to water content and the ionic environment, and should be verified using metrics such as membrane integrity and leakage rate.
(3) Discussions Related to Nucleic Acid Systems
Trehalose may influence the hydration environment and conformational stability of nucleic acid systems through hydrogen-bond networking. When stronger conclusions such as radiolysis protection are discussed, scientific statements should be bounded by clearly defined model conditions, doses, and endpoints, and supported by reproducibility validation.
IV. Preparation Methods and Process Routes
4.1 Biological Extraction
(1) Raw Material Sources
Trehalose has been identified and extracted from plant materials such as kelp, seaweeds, and aloe, and also from edible fungi such as shiitake, lichens, and maitake mycelia. In research and industrial practice, trehalose extraction and purification from yeast cells has been more widely emphasized.
(2) Typical Workflow and Assay
The 3,5-dinitrosalicylic acid (DNS) method can be used for analysis. A typical extraction workflow includes: active yeast → centrifugation → impurity removal → purification → drying.
(3) Industrialization Constraints
This traditional route is relatively mature, but its yield is often limited and the production cost and cycle time can be higher. As a result, it is constrained in large-scale modern food applications and is more often used for research, benchmarking, or specific niche scenarios.
4.2 Enzymatic Synthesis
(1) Substrates and Method Categories
Enzymatic production typically uses carbohydrates such as starch or maltose as substrates. Trehalose-related enzyme systems convert these substrates into trehalose, and processes may be classified as single-enzyme, dual-enzyme, or multi-enzyme (cocktail) routes.
(2) Dual-enzyme Route
In the dual-enzyme route, starch is used as the feedstock and conversion is achieved via coordinated action of maltooligosyltrehalose synthase and a hydrolase. In practice, the substrate is often first processed into short-chain dextrins, followed by the trehalose formation step, and then decolorization, desalting, and crystallization for downstream purification to obtain products meeting different grade requirements.
(3) Single-enzyme Route and Strain Differences
Single-enzyme routes have attracted interest due to controllable processes, relatively simplified workflows, and more manageable by-product control. Trehalose synthases from different microbial sources can differ in yield performance, and practical processes should be optimized with reaction windows and quality attributes as the guiding objectives.
(4) Key Points for Process Economics
The cost and supply stability of enzyme preparations can materially affect total production cost and should be treated as major variables in sustainability assessments of enzymatic routes. In addition, mother liquor recovery, energy consumption, and purification burden should be incorporated into overall process economic accounting.
4.3 Microbial Fermentation
Microbial fermentation is considered capable of alleviating certain process constraints associated with traditional extraction and enzymatic routes. Its core lies in accumulating trehalose or enabling trehalose conversion through strain selection and fermentation condition control, followed by extraction and purification. Key control points include carbon source strategy, osmotic pressure and stress management, dissolved oxygen and pH control, as well as downstream removal efficiency for proteins, pigments, and salt-related impurity profiles.
4.4 Genetic Engineering-based Production
Trehalose synthases are often intracellular enzymes, and wild-type strains may suffer from low enzyme activity and limited conversion efficiency. Industrial practice therefore frequently applies genetic engineering to production strains to increase synthetic flux and reduce losses through degradation pathways. Representative routes include whole-cell catalysis in Escherichia coli, one-carbon pathways in methanotrophs, photoautotrophic routes in cyanobacteria, and yeast routes that block degradation pathways. Shared engineering priorities include building stable expression systems, balancing host metabolic burden, improving product export or accumulation efficiency, and treating reaction broth complexity and downstream purification cost as equally important evaluation dimensions.
V. Pharmacology-related Research and Biological Effects
5.1 Role as a Pharmaceutical Excipient and Stabilizer for Diagnostic Reagents
In the pharmaceutical industry, trehalose is commonly used as a stabilizer for reagent-grade and diagnostic products, and as a candidate protectant or excipient in lyophilized biologics. By forming a glassy matrix, reducing molecular mobility, and supporting hydration layer stability, trehalose can suppress protein aggregation and certain deactivation pathways. This is a formulation- and process-coupled effect and should be confirmed using critical quality attributes and accelerated stability data.
5.2 Drug Delivery and Related Research Discussions
Trehalose can be used as an excipient in drug delivery systems, and exploratory research has discussed adjunctive therapeutic potential in certain disease areas. Such conclusions are typically sensitive to model type, route of administration, and exposure level. In technical and popular science materials, it is inappropriate to extrapolate preclinical findings directly into definitive human pharmacological conclusions.
5.3 Moisturization and Skin Repair-related Application Basis
Trehalose can be incorporated into moisturizing formulations and may serve as one source of energy substrate. When combined with moisturizing and film-forming components such as hyaluronic acid, trehalose can contribute to formulation systems that support the skin barrier. Statements involving specific efficacy or model data should be presented with traceable study conditions and appropriate usage boundaries.
VI. Application Fields and Technical Considerations
6.1 Food Applications
(1) Beverages (including dairy beverages, plant-protein beverages, acidic juices, etc.)
Trehalose typically does not readily engage in classic Maillard browning pathways during thermal treatment, which is beneficial for maintaining color and flavor stability. It exhibits good chemical stability from acidic to neutral formulation windows and is suitable for a wide range of beverage systems. With sweetness lower than sucrose, it can support structured design of sweetness coordination and mouthfeel roundness in reduced-sugar formulations. Functional statements such as enzyme activity stabilization should be evaluated using activity retention testing and shelf-life data under the target matrix.
(2) Baked Goods and Cereal Products
Trehalose can be used in baking systems for sweetness and solids structuring, and may support texture stability and shelf-life management under certain conditions. Its effects on browning, staling or retrogradation, and frozen dough performance are strongly influenced by formulation moisture, sugar–salt systems, and fermentation or freezing profiles, and should be evaluated via controlled studies using structure, softness, and retrogradation-related metrics.
(3) Frozen and Refrigerated Foods
In frozen systems, trehalose may support texture and mouthfeel stability by influencing freeze-concentrated phases and the tendency to form a glassy state. Its impact on ice crystal formation and recrystallization is sensitive to formulation components, freezing rate, and storage temperature fluctuations, and should be assessed using particle size distributions, melting curves, and sensory evaluation.
(4) Fried Foods
Trehalose can be used in pre-frying batter or coating systems to influence surface setting and moisture migration, thereby supporting management of oil uptake and crispness. Quantitative statements such as reduction ratios should be based on analytical data under specific product and process conditions; technical manuscripts should use appropriately conservative language.
(5) Meat and Aquatic Products
In meat and aquatic systems, trehalose can support water-holding capacity and texture stability during frozen storage, and may in certain cases inhibit lipid oxidation-related quality deterioration. Actual effects and optimal addition windows depend on formulation salinity, protein content, freeze–thaw cycles, and packaging conditions, and should be validated using drip loss, texture profile analysis, and oxidation indicators.
6.2 Pharmaceutical and Bioproduct Applications
Trehalose can be used as a stabilizer for reagent-grade and diagnostic products, and in formulation design for lyophilization and low-temperature preservation systems. Its ability to form a glassy matrix can reduce aggregation and loss of activity, but whether it meets parenteral-grade requirements depends on raw material grade, endotoxin control, microbial limits, and impurity profiles within the quality system. Statements such as plasma substitution or room-temperature preservation should be described professionally with the specific product type, applicable regulations, and validation data as boundaries.
6.3 Cosmetic Applications
Trehalose can be used as a humectant or protective component in emulsions, masks, serums, and cleansing products, and may also contribute to sweetness and sensory optimization in oral or lip products. Anhydrous trehalose can be evaluated as a dehydration-related component for phospholipid systems or enzyme-based actives in certain formulations. Practical formulation development should consider electrolyte environment, viscosity, and crystallization risk, and confirm suitability through stability testing and sensory evaluation.
6.4 Agricultural Applications
In agriculture, trehalose can be applied as an exogenous stress-resistance inducer or biostimulant via foliar spraying, seed soaking, or root drenching. Response to concentration and application frequency can differ substantially across crops and growth stages. It is recommended to rely on small-scale controlled trials to evaluate endpoints such as photosynthetic parameters, osmotic adjustment substances, antioxidant system indicators, and yield component factors. Quantitative claims regarding improvement magnitude should be presented with test conditions, sample size, and statistical analysis as prerequisites, and technical manuscripts should use conservative language.
VII. Aladdin Related Products
Catalog No. | Product Name | CAS No. | Grade or Purity |
6,6'-Diazido-6,6'-dideoxytrehalose | 18933-88-5 |
| |
D-(+)-Trehalose dihydrate | 6138-23-4 | analytical standard | |
D-(+)-Trehalose dihydrate | 6138-23-4 | suitable for plant cell culture, ≥99% | |
D-(+)-Trehalose dihydrate | 6138-23-4 | for cell culture, suitable for insect cell culture, ≥99% | |
D-(+)-Trehalose dihydrate | 6138-23-4 | 10mM in DMSO | |
D-(+)-Trehalose dihydrate | 6138-23-4 | ≥99%, from cassava starch | |
D-(+)-Trehalose Anhydrous | 99-20-7 | 10mM in DMSO | |
D-(+)-Trehalose Anhydrous | 99-20-7 | ≥99% | |
D-(+)-Trehalose-d | 2028292-31-9 | ≥99% | |
D-(+)-Trehalose-d | 1334376-67-8 |
| |
FITC -trehalose |
| average mol wt 700Da | |
α,β-Trehalose-d | 2028292-29-5 |
| |
α,β-Trehalose | 585-91-1 | ≥95% |
As a non-reducing disaccharide, trehalose features mild sweetness, good thermal and acid stability, low hygroscopicity, and a relatively high glass transition temperature, and it can provide a stabilizing environment for proteins and membrane structures in various systems. Its preparation methods have evolved from early biological extraction to predominantly enzymatic synthesis, while microbial fermentation and genetic engineering routes continue to improve process efficiency and cost structure. In practical development and evaluation, raw material form and moisture level, formulation compatibility, processing thermal history, and storage conditions should be integrated into a unified quality and validation framework to ensure rigorous and reusable conclusions.